Haloethisterone compounds



United States Patent 3,498,975 HALOETHISTERONE COMPOUNDS Arthur E.Oberster, Canton, Ohio, Roger E. Beyler, Carbondale, Ill., and Lewis H.Sarett, Princeton, N.J., assignors to Merck & Co., Inc., Rahway, N.J., acorporation of New Jersey No Drawing. Continuation-impart ofapplications Ser. Nos. 23,392, 23,393, 23,394, 23,395, 23,396, and23,397, all Apr. 20, 1960. This application Sept. 30, 1960, Ser. No.59,479

Int. Cl. C07c 167/00, 169/22; A61k 17/08 US. Cl. 260-23955 16 ClaimsABSTRACT OF THE DISCLOSURE The invention disclosed herein is concernedgenerally with novel steroids and processes of preparing the same. Moreparticularly, it relates to novel 21-halo-ethisterones,2l-halonorethisterones, and closely related compounds, which possessuseful therapeutic properties as orally and parenterally activeprogestational agents, and which are prepared by reacting thecorresponding ethisterone or 19- nor-ethisterone compound in which the3- and 17-oXygen functions are protected, with a halogenating agent.This invention also relates to pharmaceutical compositions containingthese novel steroid compounds.

This is a continuation-in-part of copending applications Ser. Nos.23,392, new Patent No. 3,246,021; 23,393, now Patent No. 3,100,204;23,394, now Patent No. 3,- 121,079; 23,395, now abandoned; 23,396, nowPatent No. 3,211,725; and 23,397, now Patent No. 3,067,214, all filedApr. 20, 1960.

In accordance with the present invention, the novel 21- haloethisterones(17 or ha1oethynl-4-androstene-l75-01-3- ones) and the2l-halonorethisterones (17ozhalo-ethyny1-3-oXygenated-androstene-1713-01 by first protecting theoxygen functions present at the C-3 and C-17 positions of the steroidmolecule with suitable protecting groups, and then halogenating to formthe 17a-haloethynyl derivative. The protecting groups at C3 and C-17 arethen removed.

This process may be schematically represented as follows, starting witha 17a-ethynyl-4-androstene-175-01-3- one in which R is methyl, or thecorresponding 19-norsteroid in which R is hydrogen:

ECH

3,498,975 Patented Mar. 3, 1970 When starting with the19-nor-17a-ethynyl-4-androstene-17fi-ol-3-one, in which R is hydrogen,Compound II-A (and likewise Compounds II-B and III) are mixtures of theA and the A -androstenes, and in the case of the Asuo) compound, thehydrogen (R) shown at C-10 is present at C-6. Removal of the protectinggroups in Step 3, using concentrated HCl gives only the S-keto-N-androstene (IV-A), but using approximately aqueous acetic acid gives amixture of the 3-keto-A (IV-A), the 3-keto-A -(IV-B) and the 3-keto-A-androstenes, which compounds can be separated by chromatography.

When starting with the 17wethynyl-4-androstene-1713- ol-3-one, in whichR is methyl, only the A -androstene is present at II-A, II-B and III,and only the M-androstene (IV-A) is formed in Step 3.

The process may be schematically represented in the following manner,starting with a 17u-ethynyl-5-androstene-3B,17;3-diol in which R ismethyl, or the corresponding 19-nor-steroid in which R is hydrogen:

HO CECE R l l l m no czox no ozox R I R i l i l l no o If the startingmaterial is the l7a-ethynyl-5-androstene- 313,17fl-diol (I'), theproduct (IV') which is obtained on removal of the protecting groups isthe l7a-haloethynyl- S-androstene-Iifi,17fl-diol. This compound may beconverted into the 17a-haloethynyll-anrostene-17;3-ol-3-one(21-haloethisterone) by oxidation, for example, with alumnumisopropoxide and cyclohexanone (or acetone) in benzene. The choice ofthe 3-oxygenated-steroid used as starting material is based on practicalconsiderations, availability and the like.

In the first step of our process the oxygen functions present at the C-3and C-17 positions of the steroid molecule are protected. In a preferredembodiment of our invention a 3-keto group is blocked by forming the 3-ethylenedioxy-derivative by reaction with excess ethylene glycol inbenzene, toluene or ethylene dichloride solvent in the presence of acidcatalysts such as p-toluenesulfonic or sulfuric acid, the waterby-product being continuously removed.

The 3-keto group may also be converted into the 3-ethylenedioxyderivative by exchange dioxolanation, which involvesacid-catlayzed transfer of the ethylene glycol portion of simple2,2-dialkyl-1,3-dioxolanes, such 2,2-dimethyl-l,3-dioxolane (acetoneethylene ketal) or, better, Z-methyl-Z-ethyl 3 dioxolane (butanoneethylene ketal), on reaction of the ketones either in an inert solvent,such as benzene, or simply by using an excess of reagent.

Other cyclic ketal derivatives can be used in our process for protectinga ketone group at C-3. In general, We have found that the loweralkylenedioxy derivatives wherein the hydrocarbon group contains notmore than seven carbon atoms, such as the ethylenedioxy,trimethylenedioxy, propylenedioxy and 'butylenedioxy derivatives aremost suitable. However, in place of using a lower alkylenedioxysubstituent to block or protect the keto substituent, We can use otherderivatives readily hydrolyzable to keto, such as an enol ethermonothioketal, or a dithioketal derivative for this purpose.

The -01 group, and the 35,17 3-di0l groups in the steroid molecule areprotected by reaction with dihydropyran in the presence of an acidicreagent, such as p-toluenesulfonyl chloride, to form the correspondingtctrahydropyranyl ether. This reaction takes place, for example, onaddition of p-toluenesulfonyl chloride to a solution of the steroid inan excess of dihydropyran. The reaction mixture is stirred at roomtemperature for 16 to 64 hours. The product may be recovered byneutralizing the reaction mixture with dilute base, and then extractingwith ether. The ether extracts are Washed with water, dried andevaporated to dryness. The crude product is dissolved in a solvent suchas petroleum ether, and chromatographed over alumina to give thecorresponding tetrahydropyranyl derivative.

In Step 2 of our invention, the steroid starting material, protected atthe 3- and the l7-position as indicated above, is treated with asuitable halogenating agent. In the preferred embodiment of ourinvention, the steroid compound, dissolved in a tertiary alcohol, isfirst treated with a potassium alcoholate of that tertiary alcohol, toform the 2l-potassium derivative of the steroid compound, and the lattercompound is then halogenated.

To form the 2l-chloro-derivative, the steroid is dissolved in t-butylalcohol and treated with potassium-tbutoxide and then witht-butyl-hypochlorite.

To form the 21-bromo-derivative, the steroid is dissolved in t-butylalcohol and treated with potassium tbutoxide and then withN-bromosuccinimide. The choice of the tertiary alcohol reagents isdetermined by practical considerations, such as availability, solubilityand the like.

The 21halo-steroid compounds are conveniently recovered by adding Waterto the reaction mixture, and extracting with ether. The ether extractsare then Washed with water, dried, and evaporated to dryness in vacuo.The residual material thus obtained is separated by chromatography onalumina to afford the 2l-halo-steroid.

In Step 3 of our process, the protecting groups at C-3 and C-l7 areremoved, preferably by treatment with acid. Removal of the protectinggroups using concentrated HCl gives the corrresponding 3-keto-A-andr0stene. To carry out this reaction, a solution of the steroid inmethanol is stirred with concentrated HCl for approximately one hour atroom temperature. The methanol may then be removed under reducedpressure. The steroid is conveniently recovered by ether extraction. Theether extract is dried, evaporated to dryness and the residual materialcrystallized from a suitable solvent.

When starting with the l9-nor-17a-ethynyl 4 androstene-l7fl-ol-3-one, inWhich R is hydrogen, removal of the protecting groups from the mixtureof the l9-nor-A and A -androstenes present in III by treatment withapproximately 70% aqueous acetic acid for about one hour at roomtemperature gives a mixture of the 3-ketol9-nor-A4-, the3-lreto-19-nor-A and the 3-keto-l9- nor-A androstenes. These compoundscan be separated by chromatography over acid-washed alumina and elutionwith mixtures of ether and petroleum ether.

The 17a-haloethynyl-5 androstene 35,175 diol (IV) may be converted intothe l7ot-haloethynyl4-androstene- 17,6-ol-3-one (2l-haloethisterone) (V)by oxidation with aluminum isopropoxide and cyclohexanone in benzene.This oxidation is carried out by dissolving the steroid in a solventsuch as a mixture of cyclohexanone and benzene and reacting in an inertatmosphere with a solution of aluminum isopropoxide in benzene at 8090C. for 2-16 hours. To recover the product, the solution is cooled, a fewdrops of water are added, and the resulting aluminum hydroxide isfiltered off. The filtrate is concentrated on the steam bath underreduced pressure, and the residual material is crystallized from asuitable solvent.

In accordance with the present invention, the novel 21- haloethisteronesare prepared from the 17a-ethynyl-5- androstene-3;8,17fl-diol by firstreacting the latter compound with dihydropyran in the presence ofp-toluenesulfonyl chloride to give the 17a-ethynyl-5-androstene-3,8,1713-diol-bis-tetrahydropyranyl ether.

The 17a ethynyl-5-androstene-3B,17B-diol-bis-tetrahydropyranyl ether ischlorinated by dissolving in t-butyl alcohol and treating with asolution of potassium t-butoxide and then with t-butyl hypochlorite togive the 170: chloroethynyl 5 androstene 3,8,17,6 diolbistetrahydropyranyl ether. The latter compound is then hydrolyzed withan acidic reagent such as concentrated HCl to give17u-chloroethynyl-5-androstene-3/3,17,8-diol, which is then oxidizedwith aluminum isopropoxide and cyclohexanone in benzene to give theflat-chloroethynyl- 4-androstene-17B-ol-3-one (21-chloroethisterone).

The 17st ethynyl-S-androstene-Elfi,l7 8-diol-bis-tetrahydropyranyl etheris brominated by dissolving the steroid in t-butyl alcohol and treatingwith a solution of potassium t-butoxide and then with N-bromosuccinimideto give the 17:1 bromoethynyl 5 androstene 35,175 diolbistetrahydropyranyl ether. The latter compound is then hydrolyzed withconcentrated HCl to give 17a-bromoethynyl-5-androstene-3 3,17fi-diol,Which is then oxidized with aluminum isopropoxide and cyclohexanone inbenzene to give the l7abromoethynyl-4-androstene-175-01- 3one(21-bromoethisterone) The novel 21-halonorethisterones are prepared fromnorethisterone (17u-ethynyl-19-nor-4-androstene-175-01-3- one) byreaction first with ethylene glycol in the presence of p-toluenesulfonicacid to give a mixture of the A and the A-17a-ethynyl-3-ethylenedioxy'19-nor-androstene- 175-01, and then Withdihydropyran to give the corresponding tetrahydropyranyl ether.

The mixture of the A and the A -l7ot-ethynyl-3-ethylenedioxy-19-norandrostene-17B-ol tetrahydropyranyl ether ischlorinated by dissolving the steroid in t-butyl alcohol and treatingWith a solution of potassium t-butox ide and then witht-butylhypochlorite to give a mixture of the A and the A-IM-chloroethynyl-l9-nor-androstene-l7fl-ol-3-one tetrahydropyranylether, which mixture is then hydrolyzed with acid. Hydrolysis of themixture with concentrated HCl gives the 17ot-chloroethynyl-19-nor-4-androstene-17/3-ol-3-one 2 1 -chloronorethisterone Hydrolysis ofthe mixture with 70% aqueous acetic acid gives a mixture of the A theA500) and the A -17u-ch1oroethynyl-19nor-androstene-17B-ol-3-one, whichmixture is separated by chromatography.

The mixture of the A and the A -17a-ethynyl-3- ethylenedioxy l9 norandrostene 17B ol tetrahydropyranyl ether is brominated by dissolvingthe steroid in t-butyl alcohol and treating with a solution of potassiumt-butoxide and then with N-bromosuccinimide to give a mixture of the Aand the A -17a-bromoethynyl-19- nor-androstene-l7B-ol-3-onetetrahydropyranyl ether. The latter compound is then hydrolyzed withacid. Hydrolysis of the mixture with concentrated HCl gives the 170:-bromoethynyl l9 nor 4 androstene-l7fi-ol-3-one (21-brornonorethisterone). Hydrolysis of the mixture with 70% aqueous aceticacid gives a mixture of the A the A and the A-17a-bromoethynyl-19-nor-androstene- 17fl-ol-3-ones, which mixture isseparated by chromatography.

In addition to the above described 17u-haloethynyl-4-androstene-l7B-ol-3-ones, this invention contemplates the preparation ofcertain derivatives thereof, and in particu- 6 lar the novel steroidcompounds which may be chemically represented as follows:

The Not-haloethynyl group of the 21-haloethisterones and the2l-halonorethisterones (VI) is hydrogenated to the correspondingchlorovinyl derivative (VI) using a Lindlar catalyst (lead deactivatedpalladium on a calcium carbonate support), and to the correspondingchloroethyl derivative (VII) using a catalyst such as platinum oxide.

VIII

The 6u-methyl-21-haloethisterone is prepared by reacting the17a-haloethynyl-5-androstene-3,8,17/i-diol with monoperphthalic acid togive the 17ot-haloethynyl-androstane-3B,l7fl-diol-5,6-oxide, which onreaction with methyl magnesium iodide yields6,8-methy1-17u-haloethynyl-androstane-3B,5a,17B-triol. The lattercompound is oxidized with an oxidizing reagent prepared from chromiumtrioxide and sulfuric acid to give 6fi-methyl-17a-haloethynyl-androstzine-5a,17/3-diol-3-one, which on treatment withaqueous sodium hydroxide gives the 60C-methyl-17a-haloethynyl-4-androstene-17fl-o1-3-one.

The 60: methyl 17a haloethynyl 4 androstene- 17B-ol-3-one isdehydrogenated at (3-1 by means of selenium dioxide, or alternately bymicrobiological methods, to give the6tx-methyl-17ot-haloethynyl-1,4-androstadiene'175-ol-3-one.

The 6oz methyl 17a haloethynyl 4 androstene- 17,8-ol-3-one is reactedwith chloranil to give 6-methy1- a haloethynyl 4,6 androstadiene 17 3 o13 one,

which compound is dehydrogenated at C-1, by means of selenium dioxide,or alternately by microbiological methods, to give6-methyl-l7oc-haloethynyl-l,4,6-androstatriene-l7p-ol-3-one.

The 17fl-lower alkyl ethers of the17B-hydroxy-17ahaloethynyl-3-keto-steroids are prepared by the reactionof the corresponding 17B-hydroxy-steroid with a lower alkyl halide andsilver oxide in a solvent such as dimethyl formamide. The lower alkylhalides which may be used for this purpose includes methyl iodide, ethyliodide, n-propyl iodide, n-butyl iodide and the like.

The l7f3-lower alkanoyl ethers of the17B-hydroxy-17ahaloethynyl-3-keto-steroids are prepared by the reactionof the corresponding 17B-hydroxy-steroid with a lower alkanoic acidanhydride in the presence of an organic base such as pyridine. The loweracid anhydrides which may be used for this purpose include aceticanhydride, propionic anhydride, butyric anhydride, and the like.

The 170: haloethynyl 1,4 androstadiene-l7B-ol-3- one acetate is preparedby reacting 17x-haloethynyl-4- androstane-l7 -ol-3-one with aceticanhydride in pyridine to give the corresponding acetate, and thenconverting the latter compound into theUna-haloethynyl-1,4-androstadiene-l7fi-ol-3-one acetate by means ofselenium dioxide, or alternately by microbiological methods.

The 170: haloethynyl 1,4 androstadiene 176- methoxy-3-one is prepared byreacting 17a-haloethynyl-4- androstene-17B-ol-3-one with methyl iodideand silver oxide, using dimethyl formamide as solvent, to give the 17ahaloethynyl 4 androstene-17B-methoxy-3-one and then converting thelatter compound into the Hot-haloethynyl-1,4-androstadiene-17B-methoxy-3-one by means of selenium dioxide, oralternately by microbiological methods.

The 60: chloro 170': haloethynyl 1,4 androstadiene-l7B-ol-3-one acetateis prepared by reacting 170chaloethynyl-4-androstene-17fl-ol-3-one withacetic anhydride and p-toluenesulfonic acid to give the 17lX-halO'ethynyl 3,5 androstadiene 3,17fl diol diacetate. The latter compound isreacted With N-chlorosuccinimide to give the6ot-chloro-17oi-haloethynyl-4-androstene-17,8-01-3- one acetate, whichis converted into the6a-chloro-171xhaloethynyl-l,4-androstadiene-17B-ol-3-one acetate byreaction with selenium dioxide, or alternately by microbiologicalmethods.

The 60; chloro 17cc haloethynyl 1,4 androstadiene-175-methoxy-3-one isprepared by reacting 17oc-halO- ethynyl-4-androstene-l7,6-ol-3-one withmethyl iodide and silver oxide, using dimethyl-formamide as solvent, togive the 17rx-haloethynyl-4-andr0stene-17/3-methoxy-3-one. The lattercompound is then converted into the l7a-ha1oethynyl 3,5androstadiened7fi-methoxy-3ol-acetate, by reaction with acetic acid andp-toluenesulfonic acid. The acetate so formed is treated withN-chlorosuccinimide to give the6a-chloro-l7u-haloethynyl-4-androstene-175-mothoxy-3-one, which compoundis dehydrogenated by selenium dioxide, or alternately by microbiologicalmethods, to the 6a-chloro-Hot-haloethynyl-1,4-androstadiene-17B-methoxy-3-One.

The 6 chloro 17a haloethynyl 4,6 androstadiene-17fl-ol-3-one acetate isprepared by reaction of 17ahaloethynyl-4-androstene -l7fi-ol3-one withchloranil to give the l7cx-haloethynyl-4,6-androstadiene-l7;8-ol-3-one(A -2l-haloethisterone), which is converted into the 170:-haloethynyl-4-androstene-175-ol-3-one-6J-oxide on reaction withperbenzoic acid. The latter compound is reacted with HCl in chloroformsolution to give the6-chloro-l7txhaloethynyl-4,6-androstadiene-l7,8-ol-3-one which forms theacetate on treatment with acetic anhydride in pyridine, and thecorresponding 17,3-methoxy-derivative on reaction with methyl iodide andsilver oxide. The 6-chlorol7ahaloethynyl-4,6-androstadiene-l7fi-ol-5-one acetate is dehydrogenated tothe 6-chloro-l7ot-haloethynyl-l,4,6-androstatriene-l7B-ol-3-one acetateon reaction with selenium dioxide, or alternately by microbiologicalmethods.

The fluoro a haloethynyl-4-androstene 17B- ol-3-one acetate is preparedfrom the Nix-haloethynyl- 3,5-androstadiene-3,175-diol diacetate byreaction high perchloryl fluoride followed by acid treatment. The 60:-fiuoro 17a haloethynyl 4 androstene ol 3- one acetate is dehydrogenatedat C1 by mean of selenium dioxide, or alternately by microbiologicalmethods, to give 600 fiuoro 17a haloethynyl 1,4 androstadiene17fi-ol-3-one acetate.

The 60: fluoro 17a haloethynyl 4 androstene- 17/3-ol-3-one acetate isreacted with chloranil to give 6 fiuoro 17cc haloethynyl 4,6androstadiene 17pol-3-one acetate, and the latter compound isdehydrogenat ed at C1 by means of selenium dioxide, or alternately bymicrobiological methods, to give6-fluoro-17e-halocthynyl-1,4,6-androstatriene-17,6-01-3-one acetate.

The 170: haloethynyl 3,5 androstadiene 175 methoxy-3,8-o1 acetate isreacted with perchloryl fluoride followed by acid treatment to give6a-fluoro-l7a-haloethynyl-4-androstene-l7fi-methoxy 3 one, whichcompound maybe dehydrogenated at C-l, by means of selenium dioxide, oralternately by microbiological methods, to give 60cfluoro-l7zx-haloethynyl 1,4 androstadiene-Ufimethoxy-3-one.

The 604 fluoro-l7a haloethynyl 4 androstenel75-methoxy-3-one isconverted into6-floro-17a-haloethynyl-4,6-androstadiene-l7,8-methoxy-3-one by the useof chloranil. The6-fluoro-l7u-haloethynyl-4,6-androstadiene-17fl-methoxy-3-one isdehydrogenated at C-1 by means of selenium dioxide, or alternately bymicrobiological methods, to yield 6-fluoro-l7a-haloethynyl-1,4,6-androstatriene-17,8-methoxy-3-one.

The l7a-haloethynyl-4-androstene-175-ol-3-one is reduced to thecorresponding 17ot-haloviny1 derivative (21-halo-4,20-pregnadiene-17/3-ol-3-one) by hydrogenation in ethyl acetate,using a Lindlar catalyst (lead deactivated palladium on a calciumcarbonate support), and to the corresponding l7ot-haloethyl derivative(21-halo-4-pregnene-176-3-one) by hydrogenation in ethyl alcohol using aplatinum oxide catalyst.

The 6a-methyl-derivatives of the 21-halonorethisterones are prepared byfirst converting the Zl-halonorethisterones (17ahaloethynyl-l9-nor-4-androstene-17,8-01-3- one) into thel7a-haloethynyl-l9-nor-5-androstene-3B, 17,8-diol-l7fl-acetate byreacting the Hot-haloethynyll9-nor-4-androstene-17;8-ol-3-one withacetic anhydride and p-toluenesulfonic acid to give the Hat-haloethynyl-19-nor-3,5-androstadiene-3,l7l3-diol-diacetate and then reacting thelatter compound with sodium borohydride. Oxidation of 17a haloethynyl 19nor-5-androstene-3fi, 17B-diol-17fl-acetate with monoperphthalic acidyields 17a haloethynyl l9 nor androstane 3,8,176 diol-S,6-oXide-17B-acetate. The latter compound is reacted with methylmagnesium iodide to form66-methyl-l7e-haloethynyl-19-nor-androstane-3B5a,17,8-triol, which onoxidation with chromic oxide in acetone gives GB-methyl-17ec-haloethynyl-19-nor-androstane-5ot,17B-diol-3-one. The lattercompound is treated with sodium hydroxide to form 60: methyl 17ozhaloethynyl-l9-nor-4-androstene-l76- ol-3-one.

The 6a-methyl-l7a-haloethynyl-l9-nor 4 androstene- 17B-ol-3-one isconverted to 6a-methyl-lWat-haloethynyl-19-nor-4-androstene-l7fi-methoxy-3-one by reaction with methyl iodideand silver oxide. The6e-methyl-17a-haloethynyl-4-androstene-17B-ol-3-one is treated withacetic anhydride in pyridine to give6tx-methyl-l7-haloethynyll9-nor-4-androstene-17,8-ol-3-one acetate.

The 6a-chloro-17e-haloethynyl-19-nor 4 androstene- 17,8-ol-3-one acetateis prepared from the Wet-haloethynyl-19-n0r-3,5-androstadiene-3,l'lfi-diol diacetate by reacting withN-chlorosuccinimide.

The 60 fluoro Uni-haloethynyl-19-nor-4-androstene- 17,3-ol-3-one acetateis prepared from the Hot-haloethynyl-19-nor-3,5-androstadiene-3,l7B-diol diacetate by reacting withperchloryl fluoride followed by acid treatment.

The 60c chloro 17a-haloethynyl-19-nor-4-androstene- 17fl-methoxy-3-oneis prepared from the 17a-haloethynyl- 19-nor-4-androstene17,8-ol-3-oneby reacting first with silver oxide and methyl iodide to form the17a-haloethynyl-19-nor-4-androsteue-17fi-methoxy-3-one. The lattercompound is then reacted with acetic anhydride and p-toluenesulfonicacid to give the 17 ot-haloethynyl-19-nor-3,5-androstadiene-I7B-methoxy-3-ol acetate, which is converted into theGot-chloro-l7ot-haloethynyl-19-nor-4- androstene-17,8-methoxy-3-one ontreatment with N-chlorosuccinimide.

The 6rx-fluoro-17a-haloethynyl-19-nor 4 androstene- 1713-methoxy-3-oneis obtained by reacting the17a-haloethynyl-19-nor-3,S-androstadiene-17fl-methoxy-3-ol acetate Withperchloryl fluoride followed by acid treatment.

The 17a-haloethynyl-l9-nor-4-androstene-17B-ol-3-one is reduced to thecorresponding 17et-halovinyl derivative(2l-halo-19-nor-4,20-pregnadiene-17/3-ol-3-one) by hydrogenation inethyl acetate, using a Lindlar catalyst (lead deactivated palladium on acalcium carbonate support), and to the corresponding 17a-haloethylderivative (21- halo-19-nor-4-pregnene-l7fi-ol-3-one) by hydrogenationin ethyl alcohol, using a platinum oxide catalyst.

The 170l-hfllOEthYIlYl-19-HOY-5 -androstene-175-ol- 3-one forms theacetate on treatment with acetic anhydride in pyridine, and thecorresponding l7fl-methoxyderivative on reaction with methyl iodide andsilver oxide. The 17a-haloethynyl-19-nor-5(10)-androstene-17B- ol-3-oneis reduced to the corresponding chlorovinyl derivative (21-halo-5( 10),20-pregnadiene-l7 8-ol 3-one) on hydrogenation with a Lindlar catalyst(lead deactivated palladium on a calcium carbonate support), and to thecorresponding chloroethyl derivative (21-halo-5(10)-pregnene-17Bol-3-one) on hydrogenation using a catalyst such as platinumoxide.

A further embodiment of our invention comprises novel pharmaceuticalcompositions containing these 21-haloethisterones and21-halonorethisterones.

The following examples illustrate methods of carrying out the presentinvention but it is to be understood that these examples are given forpurposes of illustration and not of limitation.

EXAMPLE 1 Twenty mg. of p-toluenesulfonyl chloride is added to 400 mg.of 17wethynyl-5-androstene-3fi,17B-diol in 20 ml. of dihydropyran. Theresulting mixture is allowed to stand at room temperature overnight. A2.5 N NaOH solution is added until the mixture is slightly alkaline.Water is then added and the aqueous phase extracted with 4 portions ofether, each containing approximately 50 ml. The combined ether layersare washed with water, dried over Na SO and evaporated under reducedpressure to give about 725 mg. of a non-crystalline product. The productdissolved in petroleum ether is chromatographed on 60 g. of neutralalumina and the chromatogram eluted with a 7:3 mixture of petroleumether and ether to give 400 mg. of crystalline product, the17u-ethynyl-5-androstene-3fi,17,B-diol-bis-tetrahydropyranyl ether,

LR. X232 290p A solution of about 4 grams of l7ot-E3thYI1Yl-5-3Ildl0-stene-3fl,17fi-diol-bis-tetrahydropyranyl ether in 75 ml. of t-butylalcohol is prepared. About 1.1 equivalents of a 1.0 molar potassiumt-butoxide is added and the resulting mixture refluxed for one hour,with stirring, and then cooled. About 1.84 ml. of t-butyl hypochloriteis then added in one portion and the reaction mixture is left stirringat room temperature overnight. About 100 ml. of water is added and theresulting aqueous mixture is extracted with four portions of ether, eachcontaining approximately 200 ml. The combined layers are washed withWater, dried over sodium sulfate, filtered and evaporated to dryness invacuo. The residual material is dissolved in petroleum ether andchromatographed on 120 g. of alumina. Elution with petroleum ether givesabout 10 3.10 grams (a 70% yield) of crystals of 17oc-Chl010- ethynyl 5androstene-3B,17B-diol-bis-tetrahydropyranyl ether. The crude productshows infrared peaks at 4.4a and 2.9a.

A solution of about 3 g. of the 17a-chloroethynyl-5-androstene-3B,175-diol-bis-tetrahydropyranyl ether in ml. of methanol isprepared. To this solution is added 2.5 ml. of concentrated hydrochloricacid and the reaction mixture is stirred for about 1 hour at roomtemperature. The methanol is then removed by evaporation under reducedpressure until the product crystallizes. Approximately 100 ml. of wateris then added and the resulting product is extracted with four portionsof ether, each containing about 200 ml. The combined ether extract iswashed with water, dried over sodium sulfate and evaporated to acrystalline residue. The residual crystalline material is recrystallizedseveral times from ether to give about 1.58 g. of17achloroethynyl-5-androstene-3,B,l7/3-diol which has the followingproperties, M.P. C.:

Analysis.Calculated for C H O Cl: C, 72.30; H, 8.38; Cl, 10.16. Found:C, 71.64; H, 8.63; CI. 10.48.

One hundred mg. of 17a-chloroethynyl-S-androstene- 313,17fi-di0l isdissolved in 1.0 m1. of cyclohexanone and 10 ml. of benzene in a flaskfitted with a magnetic stirrer and a reflux condenser. About 5 ml. ofthe benzene isdistilled and a stream of dry nitrogen is passed throughthe system and maintained throughout the reaction time. Then 0.5 ml. ofa 10% solution of aluminum isopropoxide in benzene is added and thereaction mixture is maintained at reflux temperature for 4 hours. Thesolution is cooled, 5 drops of water are added and the resultantaluminum hydroxide is filtered off. The filtrate is taken to drynessunder reduced pressure. The material is dissolved in ether, filtered,and the filtrate concentrated to give about 37 mg. of crudel7a-chloroethynyl-4-androstene-17B ol 3 one, M.P. 178183 C.Recrystallization from ether gives about 25 mg. of the purified product,M.P. 182l84 C. Chromatography of all mother liquors on 3 g. of aluminaand elution of the chromatogram with ether gives an additional 20 mg. ofproduct, M.P. 181184 C. Total yield 45 mg. The product has the followingproperties:

U.V. M153 241 ma, 6 15,000. LR. 2211? 2.8, 443, 6.0,. 6.18;;

Analysis.Calculated for C H O Cl: C, 72.73; H,

7.85; Cl, 10.22. Found: C, 73.41; H, 7.93; Cl, 10.81.

EXAMPLE 2 To a solution of 482 mg. of 17a-ethynyl-5-androstene-36,17B-diol-bis-tetrahydropyranyl ether in 10 ml. of tertiary-butylalcohol, is added about 1.1 equivalents of a 1.0 molar potassiumt-butoxide. The resulting mixture is refiuxed for one hour, withstirring, and then cooled. 196 mg. of N-bromosuccinimide is then addedand the reaction mixture is stirred at room temperature for about 18hours. The entire reaction mixture is dissolved in water and thenextracted with 3 portions of ether, each containing approximately 50 ml.The combined ether extracts are Washed with three portions of asaturated solution of NaHCO each portion containing approximately 25ml., then with 3 portions of water, each containing about 25 ml. Theether layer is dried over sodium sulfate, filtered and evaporated todryness. The oily residue is filtered through 20 g. of aluminum oxide togive 407 mg. of oily material which is dissolved in petroleum ether andchromatographed on 30 g. of acetone activated alkaline alumina. Elutionwith a 9:1 mixture of petroleum ether and ether yields 87 mg. of17a-bromo-ethynyl-5-androstene- 3B,17fl-diol-bis-tetrahydropyranylether. An infrared spectrum of this material shows A 4.5a.

To a solution of 400 mg. of17a-bromoethynyl-5-androstene-3,8,17,6-diol-bis-tetrahydropyranyl etherin 40 ml. of methanol is added 0.8 ml. of concentrated HCl, and thereaction mixture is stirred for one hour at room temperature. Themethanol is then removed under reduced pressure. Water is added and theresulting solution is extracted with 3 portions of ether, each portioncontaining approximately 75 ml. The combined ether extracts are washedthree times with approximately 50 ml. of water, dried over sodiumsulfate, filtered and evaporated to dryness. The residual material iscrystallized to give 230 mg. of17a-bromoethynyl-5-androstone-3t5,175-diol which has the followingproperties: M.P. 214215 C.

In. M3 3 2], 2.89, 4.55,

Analysis.Calculated for C H O Br: C, 64.10; H, 7.43; Br, 20.32. Found:C, 62.40; H, 7.65; Br, 20.50.

17a-bromoethynyl-5-androstene-3[5,17fl-diol (195 mg.) is dissolved in1.95 ml. of cyclohexanone and 20 ml. of benzene, using a flask fittedwith a magnetic stirrer and a reflux condenser. After 3 ml. of benzeneis distilled, a stream of dry nitrogen is passed through the system, andmaintained throughout the entire reaction time. After cooling to roomtemperature, there is added 0.98 ml. of a solution of aluminumisopropoxide in benzene, and the reaction mixture is refluxed for 3hours and cooled to room temperature. Ten drops of water are added andthe reaction mixture is filtered. The filtrate is taken to dryness. Theresidue is chromatographed on g. of acetone activated acid-Washedalumina and 85 mg. of product is eluted with a mixture of seven partsether to three parts petroleum ether. The17ot-bromoethynyl-4-androstene-17B- ol-3-one so obtained has thefollowing properties:

U.V. xggg 240,, 6 15,700, LR. my 2.9, 4.51, 6.0, 6.2,.

Analysis.-Calculated for C H P Br: C, 64.45; H, 6.95; Br, 20.42. Found:C, 65.16; H, 7.30; Br, 21.44.

EXAMPLE 3 To a solution of one gram of 17a-ethynyl-19-nor-4-androstene-17fl-ol-3-one dissolved in 75 ml. of benzene is added 7.5 ml.of ethylene glycol, together with 50 g. of p-toluenesulfonic acid. Thereaction mixture is heated at reflux with a water separator for hours.The reaction mixture is cooled, and about 10 ml. of bicarbonate solutionis added. The reaction mixture is extracted with 3 portions of ether,each portion containing about 100 ml. The combined extracts are washedwith water, dried over sodium sulfate and evaporated to dryness to givea mixture of the A and A -17a-ethynyl-3-ethylenedioxy-19-nor-androstene-17/3-ols.

To a solution of the total crude material from the above reaction in 10ml. of dihydropyran which has been distilled from sodium metal, is addedwith stirring, 120 mg. of p-toluenesulfonyl chloride, and the stirringis continued at room temperature for 64 hours. Saturated sodiumbicarbonate solution (10 ml.) is then added to the reaction mixture. Themixture is then extracted with 3 portions of ether, each portioncontaining about 50 ml. The extracts are combined, washed with water anddried over sodium sulfate, and evaporated to dryness in vacuo. The crudeoil (about 3.5 g.) is dissolved in petroleum ether and chromatographedon 100 g. of acetone activated alkaline alumina. The chromatogram iseluted with mixtures of ether and petroleum ether to give 1.043 g. of amixture of the A and A -l7u-ethynyl-fli-ethylenedioxy-19-nor-androstene-l7,8-ol-tetra-hydropyranyl ethers. The infraredspectrum shows bands at 2.9a and 9.0

To a stirred solution of 2.130 g. of the mixture of the A and A-17a-ethynyl-3-ethylenedioxy-19-nor-androstene-17fl-ol-tetrahydropyranylethers, dissolved in 50 ml. of dry t-butanol is added 5.5 ml. of 1 Mpotassium-tbutoxide. The reaction mixture is heated at reflux for ninetyminutes and then cooled to room temperature. Then 0.95 ml. of t-butylhypochlorite is added. The reaction mixture is left to stir at roomtemperature for about 16 hours. About 100 ml. of water is added to thereaction mixture and the aqueous solution is extracted with threeportions of ether, each portion containing about 200 ml. The extractsare combined, washed with water, dried over sodium sulfate, filtered andevaporated to dryness under reduced pressure. About 2.2 g. of crudeproduct is obtained. This is chromatographed on g. of alkaline alumina.The material dissolved in petroleum ether to which a small amount ofbenzene has been added is chromatographed on alumina and eluted withether-petroleum ether mixtures. 1.9 g. of a semi-crystalline prodnot,the 17a-chloroethynyl-3-ethylenedioxy-19-nor-5 [and5(10)]-androstene-l'lfi-ol-tetrahydropyranyl ether, is obtained.Infrared absorption shows bands at 4.45 and strong absorption in the910;r region.

To a stirred solution of 1.9 grams of 17a-chloroethynyl- 3 ethylenedioxyl9-nor-5[and 5 (10)]-androstene-l7flol-tetrahydropyranyl dissolved in 41ml. of methanol is added 3 ml. of concentrated hydrochloric acid and 2.1ml. of water. The reaction mixture is stirred at room temperature for 2/2 hours. The methanol is then removed under reduced pressure. Theresidue is dissolved in about 200 ml. of ether and washed with wateruntil the washings are neutral. The ether solution is dried over sodiumsulfate and evaporated to dryness. The crystalline residue,recrystallized from a mixture of ether and methylene chloride, gives 1.2g. of 17a-chloroethynyl-19-nor-4-androstene-17fl-ol-3-one, M.P. 194200C. A sample recrystallized for analysis from ethyl acetate has a meltingpoint of 198-201 C. and shows the following properties:

Analysis.-Calculated for C H O Cl: C, 72.20; H, 7.57; Cl, 10.65. Found:C, 72.27; H, 7.57; Cl, 9.90%.

Ten mg. of the mixture of 17a-chloroethynyl-3-ethylenedioxy A -[and M]-androstene-17,8-ol-tetrahydropyranyl ether is treated with 1 cc. of70% aqueous acetic acid for 1 hour at room temperature. The product isneutralized with aqueous sodium hydroxide and extracted with ether. Theether extract is dried and concentrated in vacuo. The crude product ischromatographed on acidwashed alumina and eluted with mixtures of etherand petroleum ether to give the l7oc-6thYI1Y1-5 10)-androstene-17fl-ol-3-one.

EXAMPLE 4 3.2 ml. of potassium t-butoxide is added to 849 mg. ofl7a-ethynyl 3 ethylenedioxy-l9-nor-5[and 5(10)]-androstene-17B-ol-tetrahydropyranyl ether, prepared as in Example3, in 28 ml. of t-butyl-alcohol. This mixture is refluxed for 75 minutesand then cooled to room temperature. 1.136 g. of N-bromosuccinimide isadded and the reaction mixture is then stirred at room temperature forabout 17 hours. The reaction mixture is added to about 60 ml. of waterand extracted with 3 portions of ether, each portion containing about 75ml. The ether extracts are combined, 'washed with 3, 50 ml. portions ofsaturated sodium bisulfite solution, and then with water until thewashings are neutral. The ether solution is dried over sodium sulfate,filtered and evaporated to dryness to yield the crude 17abromoethynyl-3-ethylenedioxy-19- nor 5 [and5(10)]-androstene-17,8-ol-tetrahydropyranyl ether which may 'be used inthe next step without purification.

The crude steroid is dissolved in 25 ml. of methanol and 0.2 ml. ofconcentrated hydrochloric acid. The mixture is stirred at roomtemperature for 2 /2 hours. The methanol is then removed under reducedpressure, water is added, and the reaction mixture is extracted with 3,75 ml. portions of ether. The ether layer is washed with water, driedover sodium sulfate, filtered and evaporated to dryness. This gives 737mg. of crude oil which is chromatographed on 44 g. of acetone activatedacidwashed alumina. Elution with a mixture of ether and petroleum ethergives a crystalline material which is recrystallized from ether to yieldabout 75 mg., MP. 180- is 182 of17a-bromoethynyl-l9-nor-4-androstene-l7fl-ol-3- one which has thefollowing properties:

U.V. A122 239 m E mol, 16,300 LR. A231? 2.85, 4.55, 6.1 and 6.2;].

Analysis.Calculated for C H O Br: C, 63.67; H, 6.68. Found: C, 64.11; H,7.05%.

Ten mg. of the mixture of 17oc-brornoethynyl-3-ethylenedioxy A -[and A]-androstene-l7fi-ol-tetrahydropyrany-l ether is treated with 1 cc. of70% aqueous acetic acid for 1 hour at room temperature. The product isneutralized with aqueous sodium hydroxide and extracted with ether. Theether extract is dried and concentrated in vacuo. The crude product ischromatographed on acidwashed alumina and eluted with mixtures of etherand petroleum ether to give the17oz-ethynyl-5(10)-androstene-17B-ol-3-one.

EXAMPLE 5 Ten grams of 17a-chloroethyny1-5-androstene-3 8,17ediol in 150ml. of tetrahydrofuran is treated with 100 ml. of 0.9 m. monoperphthalicacid in ethyl acetate and allowed to stand overnight at roomtemperature. The reaction mixture is diluted with ethyl acetate andwashed sequentially with aqueous sodium bicarbonate, sodium bisulfite,and sodium bicarbonate solution. The organic layer is dried andconcentrated in vacuo. The product is fractionally crystallized fromaqueous ethanol to yield thel7ot-chloroethynyl-androstane-313,17,B-diol-5,6a-oxide.

Five grams of l7e-chloroethynyl-androstane-3B,17B- diol-5,6e-oxide inbenzene is added to a reagent prepared from 3.4 g. of magnesium, 10 ml.of methyl iodide and 45 ml. of ether. The mixture is stirred and 70 ml.of solvent is removed by distillation. After refluxing for 5 hours, themixture is cooled, acidified with dilute hydrochloric acid and theorganic layer washed to neutrality. The organic phase is dried andconcentrated in vacuo. Crystallization from aqueous ethanol gives the6,B-methyl 17a chloroethynyl-andromane-35,5,17 8- triol.

The oxidizing reagent is prepared by diluting 2.7 g. of Cro and 2.3 ml.of concentrated sulfuric acid to 10 ml. with water. A solution of 1.90g. of 6,8-methyl- 17a-chloroethynyl-androstane-33,5u,17B-triol in 300ml. of acetone is cooled to and treated with 1.85 ml. of the oxidizingreagent. The solution is diluted with ice water and extracted withether. The ether extract is washed sequentially with water, and aqueoussodium bicarbonate and then dried and concentrated in vacuo, to give6,8-methyl 17cc chloroethynyl-andrOstane-Su,17B- diol-3-one. The crudeproduct is dissolved in 150 ml. of methanol, 75 ml. of 1 N sodiumhydroxide is add-ed and the mixture is stirred at room temperature undernitrogen for 24 hours. The mixture is concentrated to half volume invacuo at 25 C. and then poured into ice water. The product is extractedwith ether. The ether mixtures to yield 6a-methyl-17a-chloroethynyl-4-The concentrate is chromatographed on acid-washed alumina and theproduct eluted with ether-petroleum ether mixtures to yield 6u-rnethyl-17a chloroethynyl-4- androstene-17fi-ol-3-one.

In accordance with the above procedures, but starting with a17a-bromoethynyl 5 androstene-3/3,l7;3-diol in place of the6amethyl-l7a-chlyoroethynyl-4-androstene- 17 8-ol-3-one, there isobtained the 6u-methyl-l7u-bromoethynyl-4-androstene-l7r3-ol-3-one.

EXAMPLE 6 To 100 mg. of 6a-methyl-17a-chloroethynyl-4androstene-17B-ol-3-one in 5 ml. of t-butanol and 0.1 ml. of acetic acidis added 50 mg. of selenium dioxide. The mixture is refluxed undernitrogen for 18 hours; then 50 mg. of selenium dioxide is added and themixture is refluxed an additional 24 hours. The product is filtered, andthe filtrate evaporated to dryness. The residue is extracted with ethylacetate and the extract is washed successively with aqueous sodiumbicarbonate, ammonium sulfide, dilute ammonia water, water, dilutehydrochloric acid and water, and then dried over magnesium sulfate. Theextract is treated with activated charcoal and then concentrated todryness. Crystallization of the residue from a mixture of acetone andether gives 60:- methyl-lh-chloroethynyl 1,4 androstadiene-l7B-ol-3-one.

In accordance with the above procedures, but starting with the6a-rnethyl-17wbromoethynyl-4-androstenel7fi-o1-3-one in place of the6a-methyl-17a-chloroethynyl 4-androstene-17fl-ol-3-one, there isobtained the Gamethyl-lh-bromoethynyl 1,4 androstadiene-17B-ol-3- one.

Septomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., 10 mg. of 6a-methyl-17wchloroethynyl-4-androstene-l7e-ol-3-one is added to each shake flask asa dimethylformamide solution mg./ml.). After a conversion period of 24hours, the cells are removed by filtration, followed by three successiveextractions with equal volume of ethyl acetate. The extracts arecombined and concentrated in vacuo.

The 60c methyl-17a-chloroethynyl-1,4-androstadiene- 17,8-ol-3one isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA'-dehydrogenation product.

Bacillus sphaericus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10 mg. of 6a-methyl-l7achloroethynyl 4androstene-l7B-ol-3-one is added to each shake flask as adimethylformamide solution (100 mg./ml.). After a conversion period of24 hours, the cells are removed by filtration, followed by threesuccessive extractions with equal volume of ethyl acetate. The extractsare combined and concentrated in vacuo.

The 60: methyl 17a-chloroethynyl-1,4-androstadiene- 17,8-01-3-one, isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

In accordance with the above-procedures, but using theGet-methyl-17a-bromoethynyl-4-androstene-175-01-3- one in place of the6a-methyl-17a-chloroethyny1-4-androstene-17fi-o1-3-one, there isobtained the 6a-methyl-17w bromoethynyl-1,4-androstadiene--01-3-one.

EXAMPLE 7 A suspension of 1 g. of 6a-methyl-17-chloroethynyl-4-androstene-l7fl-ol-3-one and 2.4 g. of chloranil in 30 ml. of ethylacetate and 6 ml. of acetic acid is stirred and refluxed for 16 hours.The reaction mixture is cooled and filtered. The product is washedsequentially with aqueous bisulfite, aqueous potassium hydroxide andwater. The organic layer is dried and concentrated in vacuo.Chromatography yields6-methyl-17wchloroethynyl-4,6-androstadiene-l7fi-ol-3-one.

EXAMPLE 8 To 100 mg. of6-methyl-17a-chloroethynyl-4,6-androstadiene-l7fi-ol-3-one in 5 m1. oft-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxideis added and the mixture is refluxed for anadditional 24 hours. The product is filtered and the filtrate evaporatedto dryness. The residue is extracted with ethyl acetate and the extractis washed successively with aqueous sodium bicarbonate, ammoniumsulfide, dilute ammonia water, water, dilute hydrochloric acid andwater, and is then dried over magnesium sulfate. The extract is treatedwith activated charcoal and then concentrated to dryness.Crystallization of the residue from a mixture of acetone and ether gives6-methyl-l7ntchloroethynyl-l,4,6-androstatriene17fl-ol-3-one.

In accordance with the above procedures, but starting with the6-methy1-17a-bromoethynyl-4,6-androstadiene- 17fl-ol-3-one in place ofthe 6-methyl-l7a-chloroethynyl- 4,6-androstadiene-l7 8-ol-3-one there isobtained the 6- methyl 17a bromoethynyl-1,4,6-androstatriene-173-01-3-one.

Sepromyxa afiim's (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2%rEdamintSheffield Farms), glucose, and 0.5 corn steep liquor. After a 48hour incubation at 28 C., mg. of 6-methyl-17u-chloroethynyl-4,6-androstadiene-l7fl-ol-3-one is added to each shake flask as adimethylformamide solution (100 mg./ml.). After a conversion period of24 hours, the cells are removed by filtration, followed by threesuccessive extractions with equal volume of ethyl acetate. The extractsare combined and concentrated in vacuo.

The 6-methyl-17a-chloroethynyl-l,4,6 androstatriene- 17 3-ol-3-one, isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

Bacillus sphaericus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5 corn steep liquor. After a 24hour incubation at 28 C., 10 mg. of 6-methyl-17a-chloroethynyl-4,6-androstadiene 17B-0l-3-one is added to each shake fiask as adimethylformamide solution (100 mg./ml.). After a conversion period of24 hours, the cells are removed by filtration, followed by threesuccessive extractions with equal volume of ethyl acetate. The extractsare combined and concentrated in vacuo.

The 6-methyl-l7a-chloroethynyl 1,4,6-androstatriene- 17fl-ol-3-0ne, isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase, and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

In accordance with the above procedures, but starting with the6-methyl-17ot-bromoethynyl-4,G-androstadiene- 17,8-ol-3-one in place ofthe 6-methyl-l7a-chloroethynyl- 4,6-androstadiene-l7 3-ol-3-one, thereis obtained the 6- methyl-17a-bromoethynyl l,4,6-androstatriene-l7fi ol3- one.

EXAMPLE 9' A mixture of 500 mg. of the6a-methyl-l7u-chloroethynyl-4-androstene- 17,8-ol-3-one, 10 ml. ofdimethyl formamide, 20 ml. of methyl iodide, and 1.5 gms. of silveroxide is stirred at room temperature for 4 days. An additional /2 gm. ofsilver oxide is added at the end of each day. One hundred ml. ofchloroform is then added to the reaction mixture, then stirred for onehour, filtered, and the filtrate evaporated to dryness. The residual oilis chromatographed with acid-washed alumina and eluted with mixtures ofether and petroleum ether to give6tx-methyl-17ot-chloroethynyl-4-androstene-17,8 methoxy- 3-0ne.

In accordance with the above procedures, but starting with the6a-methyl-l7at-bromoethynyl-4-androstene-17,8- ol-3 one in place of the6a-methyl-l7u-chloroethynyl-4- androstene-17B-ol-3-one there is obtainedthe Got-methyl- 17oc-bromoethynyl-4-androstene-17 3-methoxy-3-0ne.

mg. of 6a-methyl-l7a-ehloroethynyll-androstenel7/3-ol-3-one is'heatedwith 1 cc. of acetic anhydride and 1.2 cc. of pyridine on the steam bathover night. The reaction mixture ;is"- then poured onto ice andextracted with chloroform. The extract is washed with Water andconcentrated. The'concentrate is chromatographed over acidwashed aluminaand eluted with mixtures of ether and petroleum ether to give6IX-II1GlIl1Yl-l70t-ChlOIOethYI1yl-4- androstene-l7B-ol-3-one acetate.

In accordance with the above procedures, but starting with the6ot-methyl-17a-bromoethynyl-4-androstene-17B- ol-3-0ne in place of the6oc-methyl-17ot-chloroethynyl-4- androstene-17fi-ol-3-one there isobtained the 6ot-methyll7ot-bromoethynyl-4-androstene-17;8-ol-3-oneacetate.

EXAMPLE 10 100 mg. of 17a-chloroethynyl-4-androstene-17 3-01-3- one(Example 1) is heated with 1 cc. of acetic anhydride and 1.2 cc. ofpyridine on the steam bath overnight. The reaction mixture is thenpoured onto ice and extracted with chloroform. The extract is washedwith water and concentrated. The concentrate is chromatographed overacid-washed alumina and eluted with mixtures of etherpetroleum and etherto give 17tX-ChlO1'O6'EhYI1Y1-4-3Hdl'0- stene 17B-ol-3-one acetate.

In accordance with the above procedure, but starting with the17ot-bromoethynyl-4-androstene-17fi-ol-3-one in place of the17m-chloroethyny1-4androstene-17,8-ol-3-0ne there is obtained the17a-bromoethynyll-androstene-1718- ol-3-one acetate.

EXAMPLE 11 To 100 mg. of 17a-chloroethynyl-4-androstene-175-01- 3-oneacetate in 5 ml. of t-butanol and 0.1 ml. of acetic acid is added-50 mg.of selenium dioxide. The mixture is refluxed under nitrogen for 18hours; then 50 mg. of selenium dioxide is added and the mixture isrefluxed an additional 24 hours. The product is filtered, and thefiltrate evaporated to dryness. The residue is extracted with ethylacetate and the extract is washed successively with aqueous sodiumbicarbonate, water, dilute hydrochloric acid and water, and then driedover magnesium sulfate. The extract is treated with activated charcoaland then concentrated to dryness. Crystallization of the residue gives17ot-c'hloroethyny1-1,4-androstadiene-1713-01- 3-one acetate.

In accordance with the above procedures, but starting with the17a-bromoethynyl 4 audrostene-17B-ol-3-one acetate in place of the17a-chloroethyny1-4-androstene- -01-3-one acetate, there is obtained the17a-bromoethynyl-1,4-androstadiene-17,8-01-3-one acetate.

Corynebacterium simplex (ATCC 6,946) is inoculated from a slant to 250ml. shake flasks containing a medium having the composition: 1 g./literyeast extract (Difco). After an 18 hour growth phase at 28 C., 10 mg. of17achloroethynyl-4-androstene-17.5-ol-3-one acetate is added to eachflask as a dimethylformamide solution (100 mg./ ml.). After a 24 hourtransformation period at 28 C., the cells are centrifuged, followed bythree ethyl acetate extracts of the cell-free broth. The extracts arecombined and concentrated in vacuo. The 17a-chloroethyny1-L4-androstadiene-17e-ol-3-one acetate is crystallized directly from theconcentrate. Paper chromatography of the product in a system utilizingformamide as the stationary phase and chloroform-benzene (1:1) as themobile phase indicates that the product possesses a polarity slightlygreater than the substrate.

Sepzomyxa affinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Shefileld Farms), 5% glucose, 0.5% corn steep liquor. After a 4-8 hourincubation at 28 C., 10 mg. of17ot-chloroethynyl-4-audrostene-1'7fl-ol-3-one acetate is added to eachshake 17 flask as a dimethylformamide solution (100 mg./ml.). After aconversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo.

The product is re-acetylated by treatment with acetic anhydride andpyridine, and then purified by recrystallization to give the17ot-chloroethynyl-1,4-androstadiene- 17B-0l-3-0ne acetate. Descendingpaper chromatography of the product in a system using formamide as thestationary phase, and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA'-dehydrogenation product.

In accordance with the above procedures, but using the17a-bromoethynyl-4-androstene-17fl-ol-3-one acetate in place of the17u-chloroethynyl-4-androstene-1718-01-3 one acetate, there is obtainedthe 17u-bromoethynyl-1,4- androstadiene-17fi-ol-3-one acetate.

EXAMPLE 1.2

A mixture of 500 mg. of the 17a-chloroethynyl-4-androstene-17/3-ol-3-one, 10 ml. of dimethylformamide, 20 ml. of methyliodide, and 1.5 gms. of silver oxide is stirred at room temperature for4 days. An additional /2 gm. of silver oxide is added at the end of eachday. One hundred ml. of chloroform is then added to the reactionmixture, which is then stirred for one hour, filtered, and the filtrateis evaporated to dryness. The residual oil is chromatographed withacid-washed alumina and eluted with mixtures of ether and petroleumether to give 17u-chloroethynyl-4-androstene-17,8-methoxy-3-one.

In accordance with the above procedures, but starting with the17wbromoethynyl-4-androstene-175-01-3-one in place of the17a-chloroethynyl-4-androstene-17fl-ol-3-one, there is obtained the17a-bromoethynyl-4-androstene-1718- methoxy-3-one.

EXAMPLE 13 To 100 mg. of 17a-chloroethynyl-4-androstene-17;3methoxy-S-one in 5 ml. of t-butanol and 0.1 ml. of acetic acid is added50 mg. of selenium dioxide. The mixture is refluxed under nitrogen for18 hours; then 50 mg. of selenium dioxide is added and the mixture isrefluxed an additional 24 hours. The product is filtered, and thefiltrate evaporated to dryness. The residue is extracted with ethylacetate and the extract is washed successively with aqueous sodiumbicarbonate, water, dilute hydrochloric acid and water, and then driedover magnesium sulfate. The extract is treated with activated charcoaland then concentrated to dryness. Crystallization of the residue from amixture of acetone and ether gives17ccchloroethynyl-l,4-androstadiene-17B-methoxy-3-one.

Septomyxa ofiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., mg. of 17a-ch1oroethynyl-4- androstene-l73-methoxy-3-one is added to each shake flask as a dimethylformamidesolution (100 mg./ml.). After a conversion period of 24 hours, the cellsare removed by filtration, followed by three successive extractions withequal volume of ethyl acetate. The extracts are combined andconcentrated in vacuo.

The 17a-chloroethynyl-1,4-androstadiene-17 3-methoxy- 3-one is readilycrystallized from the concentrate. Descending paper chromatography ofthe product is a system using formamide as the stationary phase andchloroform as the mobile phase, shows some of the steroid substrate, butis largely the somewhat more polar A'-dehydrogenation product.

Bacillus sphaerz'cus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 m1. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10 mg. of 17a-chloroethyny1-4-an- 18drostene-l7B-methoxy-3-one is added to each shake flask as adimethylformamide solution mg./ml.). After a conversion period of 24hours, the cells are removed by filtration, followed by three successiveextractions with equal volume of ethyl acetate. The extracts arecombined and concentrated in vacuo.

The 17a-chloroethynyl-1,4-androstadiene-17 8-methoxy- 3-one, is readilycrystallized from the concentrate. Descending paper chromatography ofthe product in a system using formamide as the stationary phase andchloroform as the mobile phase, shows some of the steroid substrate, butis largely the somewhat more polar A'-dehydrogenation product.

In accordance with the above procedures, but using the 170: bromoethynyl4 androstene-l75-methoxy-3-one in place of the 17cc chloroethynyl 4androstene-17/3-methoxy-3-one, there is obtained the17u-bromoethynyl-1,4- androstadiene-17,8-meth0xy-3-one EXAMPLE 14 Amixture of 1 g. of 17a-chloroethynyl-4-androstene- 17/3-o1-3-one, 10 m1.of acetic anhydride and 100 mg. of p-toluenesulfonic acid is heated onthe steam bath for one hour and allowed to stand overnight at roomtemperature. It is then poured into ice water and left to stand at roomtemperature for one half hour. The reaction mixture is then extractedwith ether. The ether extract is washed with water and aqueous sodiumbicarbonate, dried, and concentrated to yield the crude 17oc-Cl'1l0l'0-ethynyl-3,5-androstadiene-3,17B-diol diacetate. The product,crystallized from petroleum ether, has a melting point of -131 C. Theanalysis of a sample which has a melting point of -136 C. is: C, 69.75%;H, 7.23%. Calculated: C, 59.67%;H, 7.25%.

To a solution of 200 mg. of the 17a-chloroethynyl-3,5-androstadiene-3,l7/i-diol-diacetate dissolved in 2 ml. of acetic acid isadded with stirring, 56 ml. of N-chloro-succinimide and 2 ml. oftetrahydrofuran contaniing 5% of dry HCl. The reaction is stirred atroom temperature for 2 /2 hours. Sodium bicarbonate is added and thereaction mixture is extracted with ether. The ether extracts are driedand concentrated. The material is chromatographed over acid-washedalumina, and eluted with etherpetroleum ether mixtures, to give the6oc-Cl1lOI0-17a-Chl0- roethynyl-4-androstene-17B-ol-3-one acetate. Theanalytical sample is crystallized from a mixture of methylene chlorideether and has the following properties: M.P. 220230 C.

Ga chloro 17a-bromoethyny1-4-androstene-17fl-ol-3-one acetate.

EXAMPLE 15 To 100 mg. of Goachloro-17a-chloroethynyl-4-androstene-17/8-ol-3-one acetate in 5 ml. oft-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,ammonium sulfide, dilute ammonia water, water, dilute hydrochloric acidand water, and then dried over magnesium sulfate. The extract is treatedwith activated charcoal and then concentrated to dryness.Crystallization of the residue from a mixture of acetone and ether gives6a-chloro-l7otchloroethynyl-1,4-androstadiene-17B-ol-3-one acetate.

In accordance with the above procedures, but starting with theGet-chloro-17a-bromoethynyl-4-androstene-175- ol-3-one acetate in placeof the 6a-chloro-17a-chloroethynyl-4-androstene-l7B-ol-3-one acetatethere is obtained the 60t-ChlOI'D-170t bromoethynyl 1,4 androstadiene-17B-ol-3-one acetate.

Corynebacterium simplex (ATCC 6,946) is inoculated from a slant to 250ml. shake flasks containing a medium having the composition: 1 g./literyeast extract (Difco). After an 18 hour growth phase at 28 C., 10 mg. of6mchlorol7a chloroethynyl 4 androstene-l7fi-ol-3-one acetate is added toeach flask as a dimethylformamide solution (100 mg./ml.). After a 24hour transformation period at 28 C., the cells are centrifuged, followedby three ethyl acetate extracts of the cell-free broth. The extracts arecombined and concentrated in vacuo. The 60:-chlor-17u-chloroethynyl-1,4-androstadiene l7fl-ol- 3- one acetate iscrystallized directly from the concentrate. Paper chromatography of theproduct is a system utilizing formamide as the stationary phase andchloroform-benzene (1:1) as the mobile phase indicates that the productpossesses a polarity slightly greater than the substrate.

Septomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), glucose, and 0.5% corn steep liquor. After a 48 hourincubation at 28 C., mg. of6a-chloro-17a-chloroethynyl-4-androstene-l7B-ol-3-one acetate is addedto each shake flask as a dimethylformamide solution (100 mg./ ml.).After a conversion period of 24 hours, the cells are removed byfiltration, followed by three successive extractions with equal volumeof ethyl acetate. The extracts are combined and concentrated in vacuo.

The product is re-acetylated by treatment with acetic anhydride andpyridine, and then purified by recrystallization to give the6a-chloro-17a-chloroethynyl-l,4-androstadiene-l7fl-ol-3-one acetate, isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

In accordance with the above procedures, but using the6a-chloro-Nix-bromoethynyl 4 androstene-17/3-ol-3-one acetate in placeof the 61x chloro-l7a-chloroethynyl-4- androstene 17fi-ol-3-one acetate,there is obtained the60schloro-17a-brornoethynyl-1,4-androstadiene-17B-ol-3 one acetate.

EXAMPLE 16 A mixture of 1 g. of l7ot-chloroethynyl-4-androstene-17B-methoxy-3-one (Example 10), 10 ml. of acetic anhydride and 100 mg.of p-toluene sulfonic acid is heated on the steam bath for one hour andallowed to stand overnight at room temperature. It is then poured intoice water and left to stand at room temperature for one half hour. Thereaction mixture is then extracted with ether. The ether extract iswashed with water and aqueous sodium bicarbonate, dried, andconcentrated to yield thel7a-cl1loroethynyl-3,S-androstadiene-l7,8-methoxy-3-ol acetate.

To a solution of 200 mg. of the 17a-chloroethynyl-3,5-andr-ostadiene-l7B-methoxy-3-ol-acetate dissolved in 2 ml. of aceticacid is added, with stirring, 56 ml. of N- chlorosuccinmide and 2 ml. oftetrahydrofuran containing 5% of dry I-ICl. The reaction is stirred atroom temperature for 2 /2 hours. Sodium bicarbonate is added. Thereaction mixture is then extracted with ether. The ether extracts aredried and concentrated. The material is chromatographed over acid-washedalumina, and eluted with ether-petroleum ether mixtures to give6a-chloro17t3- chloroethynyl-4-androstene-l7fl-methoxy-3-one.

In accordance with the above procedures, but starting with the17a-bromoethynyl-4-androstene 17,8-methoxy-3- one in place of thel7a-chloroethynyl-4-androstene-17B- methoxy-3-one, there is obtained asan intermediate, the 17a-bromoethynyl-3,S-androstadiene-17B-methoxy-3-olacetate, and as product, the6a-bromo-Hoe-chloroethynyl 4-androstene-17fi-methoxy-3 -one.

EXAMPLE 17 To mg. of l7a-chloroethynyl-4-androstene-l71imethoxy-B-one in5 ml. of t-butanol and 0.1 ml. of acetic acid is added 50 mg. ofselenium dioxide. The mixture is refluxed under nitrogen for 18 hours;then 50 mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,ammonium sulfide, dilute ammonia water, Water, dilute hydrochloric acidand Water, and then dried over magnesium sulfate. The extract is treatedwith activated charcoal and then concentrated to dryness.Crystallization of the residue from a mixture of acetone and ether givesNix-chloroethynyl-1,4-androstadiene-l7fimethoxy-3-one.

Septomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., 10 mg. of17a-chloroethynyl-4-androstene-l7fi-rnethoxy-3-one is added to eachshake flask as a dimethylformamide solution (100 ing/ml). After aconversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo.

The chloroethynyl 1,4 androstadiene methoxy-3-one is readilycrystallized from the concentrate. Descending paper chromatography ofthe product in a system using formamide as the stationary phase andchloroform as the mobile phase, shows some of the steroid substrate, butis largely the somewhat more polar A'- dehydrogenation product.

Bacillus sphaerzcus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50- ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10 mg. ofl7a-chloroethynyll-androstene-l7fl-methoxy 3-one is added to each shakeflask as a dimethylformamide solution (100 mg./ml.). After a conversionperiod of 24 hours, the cells are removed by filtration, followed bythree successive extractions with equal volume of ethyl acetate. Theextracts are combined and concentrated in vacuo.

The l7a-chloroethynyl-l,4-androstadiene-17B-methoxy- 3-one is readilycrystallized from the concentrate. Descending paper chromatography ofthe product in a system using formamide as the stationary phase andchloroform as the mobile phase, shows some of the steroid substrate, butis largely the somewhat more polar A-dehydrogenation product.

In accordance with the above procedures, but using the 170: bromoethynyl4 androstene 17B methoxy 3- one in place of thel7a-chloroethynyl-4-androstene-17B- methoxy-3-one, there is obtained theNot-bromoethynyl.- l,4-androstadiene-17B-methoxy-3-one.

EXAMPLE 18 A suspension of l g. of 17u-chloroethynyl-4-androstene-17fl-ol-3-one and 2.4 g. of chloranil in 30 ml. of ethyl acetate and 6ml. of acetic acid is stirred and refluxed for 16 hours. The reactionmixture is cooled and filtered. The product is washed sequentially withaqueous bisulfite, aqueous potassium hydroxide and water. Chromatographyyields 17a chloroethynyl-4,6-androstadiene-17fl-ol-3-one which oncrystallization from ethyl acetate has the following properties: M.P.2092l1 C.: oc l59 (C 0.9 CHCI U.V. A232? 283 mu 6 26900 Calculated for CH O Cl: C, 73.l5; H, 7.31. Found: C, 73.08; H, 7.06.

A solution of 675 mg. of17u-chloroethynyl-4,6-androstadiene-l7B-ol-3-one in 30 ml. of ether and30 ml. of 0.30 N perbenzoic acid in benzene is allowed to stand in thedark for 68 hours. The solution is washed sequentially with aqueoussodium bisulfite, water, 2.5 N potassium hydroxide and water. Theorganic layer is dried and concentrated in vacuo. Chromatography onacid-washed alumina and elution with ether affords about 180 mg. of the17m chloroethynyl 4 androstene-l7;8-ol-3-one-6,7- oxide, which has thefollowing properties: M.P. 196- 216 C.

U.V. A252 240 my, e 377 A solution of 140 mg. of the oxide in 10 ml. of0.4 N hydrogen chloride in chloroform is allowed to stand at roomtemperature for 5.5 hours. The solution is poured onto ice. Aqueoussodium bicarbonate solution is added and the product is extracted withchloroform. The organic layer is washed with water, dried andconcentrated. Crystallization from ether affords 84 mg. of6-chloro-l7achloroethynyl-4,6-androstadiene-17,8-ol-3-one, M.P. 195- 205C. The sample for analysis is crystallized from methanol and has thefollowing properties: M.P. 203208 C.; a 96 (C 1.0 CHCl U.V. M1552 285 m6 21,400

Calculated for C H O Cl C, 66.49; H, 6.38. Found: C, 66.28; H, 6.62.

In accordance with the above procedures, but starting with the17a-bromoethynyl-4-androstene-17B-ol-3-one in place of the17a-chloroethynyl-4-androstene-l7fi-ol-3-one, there is obtained asintermediate, the l7a-brornoethynyl-4-androstene-l7/3-ol-3-one-6,7-oxide, and as product, the 6- chloro 17ccbromoethynyl 4,6 androstadiene 175- ol-3-one.

A solution of 125 mg. of 6-chloro-17a-ch1oro-ethynyl-4,6-androstadiene-l7 S-ol-3-one in 1.8 ml. of pyridine and 1.5 ml. ofacetic anhydride is heated on the steam bath overnight. The solution ispoured into ice water and extracted with ether. The ether solution iswashed sequentially with dilute hydrochloric acid, water and sodiumbicarbonate solution. Chromatography on 7.0 g. of acidwashed alumina,and elution with ether; petroleum ether mixtures aifords 56 mg. of6'chloro-Una-chloroethynyl- 4,6-androstadiene-175-ol-3-one acetate. Thesample for analysis is crystallized from ether and has the followingproperties: M.P. 203-208 C., a -98 (C 0.9 CHC U.V. A35 284 m 6 21,300Calculated for C H O Cl C, 65.57; H, 6.21. Found: C, 66.06; H, 6.16.

In accordance with the above procedures, but starting with the 6chloro-17a-bromoethynyl-4,6-androstadiene- 17/3-ol-3-one in place of the6-chloro-l7a-chloroethynyl- 4,6-androstadiene-l7fl-ol-3-one, there isobtained the 6- chloro 17oz bromoethynyl-4,6-androstadiene-17,8-01-3-one acetate.

EXAMPLE 19 To 100 mg. of 6-chloro-17a-chloroethynyl-4,6-androstadiene-17f3-ol-3-one acetate in 5 ml. of t-butanol and 0.1 ml. ofacetic acid is added 50 mg. of selenium dioxide. The mixture is refluxedunder nitrogen for 18 hours; then 50 mg. of selenium dioxide is addedand the mixture is refluxed an additional 24 hours. The product isfiltered, and the filtrate evaporated to dryness. The residue isextracted with ethyl acetate and the extract is washed successively withaqueous sodium bicarbonate, water, dilute hydrochloric acid and water,and then dried over magnesium sulfate. The extract is treated withactivated charcoal and then concentrated to dryness. Crystallization ofthe residue gives 6 chloro 17oz chloroethynyl-1,4,6- androstatriene-l7S-ol-3-one acetate.

Coryneb-acterium simplex (ATCC 6,946) is inoculated from a slant to 250ml. shake flasks containing a medium having the composition: 1 g./literyeast extract (Difco).

After an 18 hour growth phase at 28 C., 10 mg. of 6- chloro 17achloroethynyl 4,6 androstadiene 17 3 ol-3-one acetate is added to eachflask as a dimethylformamide solution mg./ml.). After a 24 hourtransformation period at 28 C., the cells are centrifuged, followed bythree ethyl acetate extracts of the cell-free broth. The extracts arecombined and concentrated in vacuo. The 6 chloro 17a chloroethynyl 1,4,6androstratriene-17/3-ol-3-one acetate is crystallized directly from theconcentrate. Paper chromatography of the product in a system utilizingformamide as the stationary phase and chloroform-benzene (1:1) as themobile phase indicates that the product possesses a polarity slightlygreater than the substrate.

Sepzomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., 10 mg. of 6 chloro 17achloroethynyl-4,6-androstadiene-l7fl-ol-3-one acetate is added to eachshake flask as a dimethylformamide solution (100 mg./ml.). After aconversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo.

The product is re-acetylated by treatment with acetic anhydride andpyridine, and then purified by recrystallization to give the6-chloro-Nix-chloroethynyl-1,4,6-androstatriene-17B-ol-3-one acetate.Descending paper chromatography of the product in a system usingformamide as the stationary phase and chloroform as the mobile phase,shows some of the steroid substrate, but is largely the somewhat morepolar A-dehydrogenation product.

In accordance with the above procedures, but using the 6 chloro 1704bromoethynyl 4,6-androstadiene-17fiol-3-one in place of the6-chloro-17a-chloroethynyl-4,6- androstadiene-l7fl-ol 3 one, there isobtained the 6- chloro 17a bromoethynyl 1,4,6 androstatriene-Ufiol-3-oneacetate.

EXAMPLE 20 A mixture of 500 mg. of 6-chloro-Not-chloroethynyl-4,6-androstadiene-l7fi-o1-3-one, 10 ml. of dimethylformamide, 20 ml. ofmethyl iodide, and 1.5 gins. of silver oxide is stirred at roomtemperature for 4 days. An additional /2 gm. of silver oxide is added atthe end of each day. 100 ml. of chloroform is then added to the reactionmixture which is then stirred for one hour, filtered. The filtrate isthen taken to dryness. The oil is chromatographed with acid-washedalumina and eluted with mixtures of ether and petroleum ether to givethe 6-chloro- 17a-chloroethynyl-l7,3-methoxy-4,6-androstadiene-3-one.

In accordance with the above procedures, but starting with the 6chloro-17a-bromoethynyl-4,6-androstadiene- 17,8-ol-3-one in place of the6-chloro-Hot-chloroethynyl- 4,6-andr0stadiene-l7fl-ol-3-0ne there isobtained the 6 chloro 17cc bromoethynyl 17,8 methoxy 4,6androstadiene-3-one.

EXAMPLE 21 To 100 mg. of6a-chloro-17a-chloroethyny1-17fl-methoxy-4,6-androstadiene-3-one in 5ml. of t-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,ammonium sulfide, dilute ammonia water, water, dilute hydrochloric acidand water, and then dried over magnesium sulfate. The extract is treatedwith activated charcoal and then concentrated to dryness.Crystallization of the residue from a mixture of acetone and ether givesthe 6-chloro- 17oz chloroethynyl 17B methoxy 1,4,6 androstatriene-3one.

Septomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., mg. of 60c-ChlOfO-17ot-Chl0l0-ethynyl-l7B-methoxy-4,6-androstadiene-3-one is added to each shake flaskas a dimethylformamide solution (100 mg./rnl.). After a conversionperiod of 24 hours, the cells are removed by filtration, followed bythree successive extractions with equal volume of ethyl acetate. Theextracts are combined and concentrated in vacuo.

The '6-chloro-l7a-chloroethynyl-17B methoxy 1,4,6- androstatriene-3-oneis readily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA'-dehydrogenation product.

Bacillus sphaericus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10 mg. of6-chloro-17a-chl0roethynyl-4,fi-androstadiene-17,8 methoxy 3 one isadded to each shake flask as a dimethylformamide solution (100 mg./ml.).After a conversion period of 24 hours, the cells are removed byfiltration, followed by three successive extractions with equal volumeof ethyl acetate. The ex tracts are combined and concentrated in vacuo.

The 6-chloro-l7a-chloroethynyl 175 methoxy 1,4,6- androstatriene 3 oneis readily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

In accordance with the above procedures, but using the6-chloro-17a-bromoethynyl 4,6 androstadiene 17/3- methoxy-3-one in placeof the 6-chloro-17a-chloroethynyl- 4,6-androstadiene 17 8 methoxy 3 one,there is obtained the 6-chloro-17a-bromoethyny1 1,4,6 androstatrienel7/3-methoxy-3 -one.

EXAMPLE 22 A solution of the 17a-chloroethynyl-3,S-androstadiene-3;.8,17-diol diacetate (Example 10) in aqueous tetrahydrofuran istreated with a slow stream of perchloryl fluoride for one hour and thenallowed to stand at room temperature for an additional 2 hours. Thereaction mixture is poured on ice and extracted with chloroform. Thechloroform layer is washed with aqueous bicarbonate solution, dried andconcentrated in vacuo. The crude material is dissolved in acetic acidsaturated with gaseous hydrogen chloride and allowed to stand for onehour at room temperature. The reaction mixture is poured on ice andextracted with chloroform. The chloroform layer is washed with aqueousbicarbonate solution, dried and concentrated in vacuo. The concentrateis chromatographed on acidwashed alumina to yield the6a-fiuoro-17a-chloroethynyl- 4-androstene-17fl-ol-3-one acetate.

In accordance with the above procedures, but starting with thel7a-bromoethynyl-3,5 androstadiene 313, 1713- diol diagetate in place ofthe l7a-chloroethynyl-3,5-androstadiene-318,l7B-diol diacetate there isobtained the 6a-fluoro-l7a-bromoethynyl 4 androstene 175 ol- 3-oneacetate.

EXAMPLE 23 To 100 mg. of6a-fluoro-17a-chloroethynyl-4-androstene-l7fi-ol-3-one acetate in 5 ml.of t-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,water, dilute hydrochloric acid and water, and then dried over magnesiumsulfate. The extract is treated with activated charcoal and thenconcentrated to dryness. Crystallization of the residue from a mixtureof acetone and ether gives Got-fluoro-lh-chloroethynyl-1,4-androstadiene17,8 ol 3 one acetate.

Corynebwcterium simplex (ATCC 6,946) is inoculated from a slant to 250ml. shake flasks containing a medium having the composition: 1 g./literyeast extract (Difco). After an 18 hour growth phase at 28 C., 10 mg. of6a-tluoro-17a-chloroethynyl-4-androstene-175-01 3 one acetate is addedto each flask as a dimethylforrnarnide solution mg./ml.). After a 24hour transformation period at 28 C., the cells are centrifuged, followedby three ethyl acetate extracts of the cell-free broth. The extracts arecombined and concentrated in vacuo. The'6tl-fll10I'O-170L-Ch1OI'Oethyl'ly1 1,4 androstadiene 17,8- ol-3-oneacetate is crystallized directly from the concentrate. Paperchromatography of the product is a system utilizing formamide as thestationary and chloroform-benzene (1:1) as the mobile phase indicatesthat the product possesses a polarity slightly greater than thesubstrate.

Septomyxa afiinis( ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., 10 mg. of fioc-fiUOrO-l'la-ChlOIO-ethynyl-4-androstene-17/3-ol-3-one acetate is added to each shake flaskas a dirnethylformamide solution (100 mg./ ml). After a conversionperiod of 24 hours, the cells are removed by filtration, followed bythree successive extractions with equal volume of ethyl acetate. Theextracts are combined and concentrated in vacuo.

The product is re-acetylated by treatment with acetic anhydride andpyridine, and then purified by recrystallization to give the6cx-flll0r0-17zx-Chl0l06thYl1Yl-L4- androstadiene-l7/3-ol-3-one acetate.Descending paper chromatography of the product in a system usingformarnide as the stationary phase and chloroform as the mobile phase,shows some of the steroid substrate, but is largely the somewhat morepolar A-dehydrogenation product.

In accordance with the above procedures, but using the 60:fluoro-17a-bromoethynyl-4-androstene-1713-01-3- one acetate in place ofthe 6a-fluoro-l7u-chloroethynyl- 4-andr0stene-l7B-ol-3-one acetate thereis obtained the 6a-fluoro-17e-bromoethynyl-1,4-androstadiene ol- 3-oneacetate.

EXAMPLE 24 A suspension of 1 g. of Got-fluoro-17a-chloroethynyl-4-androstene-17fl-ol-3-one acetate 2.4 g. chloranil, 30 ml. ethyl acetateand 6 ml. of acetic acid is stirred and refluxed for 16 hours. Thereaction mixture is cooled and filtered. The product is washedsequentially with aqueous bisulfite, aqueous potassium hydroxide andwater. The organic layer is dried and concentrated in vacuo.Chromatography yields6-fluoro-17u-chloroethynyl-4,fi-androstadiene-l7fi-ol-3-one acetate.

In accordance with the above procedures, but starting with theGot-fluoro-17a-bromoethynyl-4-androstene-17B- ol-3-one acetate in placeof the 6a-fluoro-17u-chloro-ethynyl-4-androstene-17B-ol-3-one acetatethere is obtained the 6 fluoro-l7ot-bromoethynyl-4,fi-androstadiene 17aol-3-one acetate.

EXAMPLE 25 To 100 mg. of6-fluoro-17a-chloroethynyl-4-6-androstadiene-17fl-ol-3-one acetate in 5ml. of t-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,Water, dilute hydrochloric acid and water, and then dried over magnesiumsulfate. The extract is treated with activated charcoal and thenconcentrated to dryness. Crystallization of the residue gives6-fiuoro-Not-chloroethynyl- 1,4,6-androstatriene-17/3-ol-3-one.

In accordance with the above procedures, but starting with the 6fluoro-17u-bromoethynyl-4,6-androstadiene- 17B-o1-3-one acetate in placeof the 6-fluoro-17a-chloroethynyl-4,6-androstadiene-17B ol-3-oneacetate, there is obtained the6-fluoro-17ut-bromoethynyl-1,4,6-androstatriene-17B-ol-3-one acetate.

Coryrzebacterium simplex (ATCC 6,946) is inoculated fr m a slant to 250ml. shake flasks containing a medium having the composition: 1 g./literyeast extract (Difco). After an 18 hour growth phase at 28 C., 10 mg. of6- fluoro-17ot-chloroethynyl-4,6-androstadiene 175-ol-3-one acetate isadded to each flask as a dimethylformamide solution (100 mg./ml.). Aftera 24 hour transformation period at 28 C., the cells are centrifuged,followed by three ethyl acetate extracts of the ce1l-free broth. Theextracts are combined and concentrated in vacuo. The 6-fluoro-17ct-chloroethynyl-1,4,6-androstatriene 175 ol-3- one acetate iscrystallized directly from the concentrate. Paper chromatography of theproduct in a system utilizing formamide as the stationary phase andchloroformbenzene (1:1) as the mobile phase indicates that the productpossesses a polarity slightly greater than the substrate.

Septomyxa afiinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., mg. of 6-fluoro-l7ot-chloroethynyl-4,6-androstadiene-17B-ol-3-one acetate is added to each shake flask as adimethylformamide solution (100 mg./ml.). After a conversion period of24 hours, the cells are removed by filtration, followed by threesuccessive extractions with equal volume of ethyl acetate. The extractsare combined and concentrated in vacuo.

The product is re-acetylated by treatment with acetic anhydride andpyridine, and then purified by recrystallization to give the6-fluoro-Uni-chloroethynyl-1,4,6- androstatriene-l7B-ol-3-one acetate.Descending paper chromatography of the product in a system usingformamide as the stationary phase and chloroform as the mobile phase,shows some of the steroid substrate, but is largely the somewhat morepolar A'-dehydrogenation product.

In accordance With the above procedures, but using the6-fluoro-17a-bromoethynyl-4,6-androstadiene 17B ol-3- one acetate inplace of the 6-fluoro-17a-chloroethynyl- 4,6-androstadiene-l7 3-ol-3-oneacetate, there is obtained the 6fluoro-17a-bromoethynyl-1,4,6androstatriene-17,3- ol-3-one acetate.

EXAMPLE 26 A solution of the 17a-chloroethynyl-3,5-androstadiene-17/i-methoxy-3fl-ol in aqueous tetrahydrofuran is treated with a slowstream of perchloryl fluoride for one hour and then allowed to stand atroom temperature for an additional 2 hours. The reaction mixture ispoured on ice and extracted with chloroform. The chloroform layer iswashed with aqueous bicarbonate solution, dried and concentrated invacuo. The crude material is dissolved in acetic acid saturated withgaseous hydrogen chloride and allowed to stand for one hour at roomtemperature. The reaction mixture is poured on ice and extracted withchloroform. The chloroform layer is washed with aqueous bicarbonatesolution, dried and concentrated in vacuo. The concentrate ischromatographed on acid-washed alumina to yield the6a-fluoro-17a-chloroethynyl-4-androstene'175-methoxy8one.

In accordance with the above procedures, but starting 26 with the17a-br0moethynyl-3,S-androstadiene 175 methoxy-3B-ol in place of the17a-chloroethynyl-3,5-androstadiene-17B-methoxy-3fl-ol, there isobtained the 6stfluoro-17u-bromoethynyl-4-androstene 17,8 methoxy-3-one.

EXAMPLE 27 To mg. of6a-fluoro-17a-chloroethynyl-4-androstene-17fi-methoxy-3-one in 5 ml. oft-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicarbonate,wjater, dilute hydrochloric acid and water, and then dried overmagnesium sulfate. The extract is treated with activated charcoal andthen concentrated to dryness. Crystallization of the residue gives6a-fiuoro-17a-chloroethynyl-1,4-androstadiene-17fl-methoxy-3-one.

Septomyxa aflinis (ATCC 6,737) is inoculated from a slant into 250 m1.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 20 C., 10 mg. of6u-fluoro-17a-chloroethynyl-4-androstene-I7B-methoxy-3-one is added toeach shake flask as a dimethylformamide solution (100 mg./ ml.). After aconversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo.

The 6ot-fluoro-17a-chloroethynl-1,4-androstadiene-l7/3- methoxy-3-one isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

Bacillus sphaericus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheffield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10' mg. of 6ot-fil101'O-170L-Chl0l'0-ethynyl-4-androstene-l7t3-methoxy-3-one is added to each shake flask asa dimethylformamide solution (100 mg./ ml.). After a conversion periodof 24 hours, the cells are removed by filtration, followed by threesuccessive extractions with equal volume of ethyl acetate. The extractsare combined and concentrated in vacuo.

The 6oz-fiUOl0-l7oc chloroethynyl 1,4 androstadiene- 17B-methoxy-3-oneis readily crystallized from the coucentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

In accordance with the above procedures, but using the 60: fluoro 17abromoethynyl 4 androstene 17B- methoxy-3-one in place of the6ct-fltl01'O-l7m-Chl0f0- ethynyl-4-androstene-17B-methoxy-3-one, thereis obtained the 6a-fluoro-l7ot-bromoethynyl-1,4-androstadiene-17B-methoxy-3-one.

EXAMPLE 28 A suspension of 1 g. of Got-fluoro-Not-chloroethynyl-4-androstene-17,8-methoxy-3-one, 2.4 g. chloranil, 30 ml. ethyl acetateand 6 ml. of acetic acid is stirred and refluxed for l6-hours. Thereaction mixture is cooled and filtered. The product is washedsequentially with aqueous bisulfite, aqueous potassium hydroxide andwater. The organic layer is dried and concentrated in vacuo.Chromatography yields6-fluoro-17u-chloroethynyl-4,6-androstadiene-17fi-methoxy-3-one.

27 EXAMPLE 29 To 100 mg. of6-fluoro-17a-chloroethynyl-4,6-androstadiene-17fi-methoxy-3-one in ml.of t-butanol and 0.1 ml. of acetic acid is added 50 mg. of seleniumdioxide. The mixture is refluxed under nitrogen for 18 hours; then 50mg. of selenium dioxide is added and the mixture is refluxed anadditional 24 hours. The product is filtered, and the filtrateevaporated to dryness. The residue is extracted with ethyl acetate andthe extract is washed successively with aqueous sodium bicardonate,water, dilute hydrochloric acid and water, and then dried over magnesiumsulfate. The extract is treated with activated charcoal and thenconcentrated to dryness. Crystallization of the residue gives6-fluoro-17a-chloroethynyl-1,4,6- androstatriene-17B-methoxy-3-one.

Seplomyxa affinis (ATCC 6,737) is inoculated from a slant into 250 ml.shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 48hour incubation at 28 C., mg. of6-fluoro-17u-chloroethynyl-4,6-androstadiene-17p-methoxy-3-one is addedto each shake flask as a dimethylformamide solution (100 mg./ml.). Aftera conversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo;

The 6-fluor0-l7a-chloroethynyl 1,4,6 androstatrienel7B-methoxy-3-one isreadily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA-dehydrogenation product.

Bacillus sphaericus (ATCC 12,488) is inoculated from a slant into 250ml. shake flasks containing 50 ml. of the following medium: 2% Edamin(Sheflield Farms), 5% glucose, and 0.5% corn steep liquor. After a 24hour incubation at 28 C., 10 mg. of6-fluoro-l7a-chlorothynyl-4,6-androstadiene-17/i-methoxy-3-one is addedto each shake flask as a dimethylformamide solution (100 mg./ml.). Aftera conversion period of 24 hours, the cells are removed by filtration,followed by three successive extractions with equal volume of ethylacetate. The extracts are combined and concentrated in vacuo.

The 6 fluoro-17u-chloroethynyl-1,4,6-androstatriene- 17B-methoxy-3-oneis readily crystallized from the concentrate. Descending paperchromatography of the product in a system using formamide as thestationary phase and chloroform as the mobile phase, shows some of thesteroid substrate, but is largely the somewhat more polarA'-dehydrogenation product.

In accordance with the above procedure, but using the 6 fluoro 170abromoethynyl 4,6-androstadiene-l75- methoxy-3-one in place of the6-fiuoro-17a-chloroethynyl- 4,6 androstadiene-17fl-methoxy-3-one, thereis obtained the 6 fluoro-17a-bromoethynyl-1,4,5,6 androstatriene-17fl-methoxy-3-one.

EXAMPLE 30 A solution of 100 mg. of the17a-chloroethynyl-4-androstene-17B-ol-3-one and 50 mg. of Lindlarcatalyst (lead deactivated palladium on a calcium carbonate support) in10 cc. of ethyl acetate is treated with hydrogen until one mole ofhydrogen has been absorbed. The mixture is filtered and concentrated.Chromatography yields the corresponding 17o: chlorovinyl derivative(21-chloro-4,20- pregnadiene-17B-ol-3-one).

In accordance with the above procedures, but starting with the 170:bromoethynyl-4-androstene-17B-ol-3-one in place of the 1701.chloroethynyl-4-androstene-l7 8-ol-3-one there is obtained thecorresponding 17u-bromovinyl derivative(21-bromo-4,20-pregnadiene-17fi-ol-3-one).

A suspension of platinum oxide in 10 cc. of ethanol is reduced and 100mg. of 17a-chloroethynyl-4-androstene- 17fl-ol-3-one is added. Reductionproceeds until two moles of hydrogen have been absorbed. The solution isfiltered, concentrated and chromatographed on alumina to yield thecorresponding 17:: chloroethyl derivative (21-chloro-4-pregnene-17B-ol-3-one) In accordance with the above procedures, butstarting with the 170: bromoethynyl-4-androstene-17fl-ol-3-one in placeof the 170; chloroethynyl-4-androstene-17 8-01-3- one there is obtainedthe corresponding 17a bromoethyl derivative(2l-bromo-4-pregnene-17B-ol-3-one).

EXAMPLE 31 A mixture of 1 g. 17 chloroethynyl 191x nor 4-androstene-17p-ol-3-one, 10 ml. of acetic anhydride and 100 mg. ofp-toluenesulfonic acid is heated on the steam bath for one hour andallowed to stand overnight at room temperature. It is then poured intoice water and left to stand at room temperature for one half hour. Thereaction mixture is then extracted with ether. The ether extract iswashed with water and aqueous sodium bicarbonate, dried, andconcentrated to yield the crude17a-chloroethynyl-19-nor-3,5-androstadiene 3,175 diol diacetate.

A solution of 8 gms. of 170: chloroethynyl-l9-nor-3,5,-androstadiene-3,l7-diol acetate in a mixture of 700 ml. of ethanol and300 ml. of tetrahydrofuran is cooled to 10 C. and added dropwise, withoccasional stirring, over a one hour period, to a cold (0 C.) solutionof 18 g. sodium borohydride in 400 ml. of 80% ethanol, the reactiontemperature not being allowed to exceed 5 C. After completion of theaddition, the solution is held at O-S" C. for an additional two hours. Asolution of NaH PO is then added to adjust the pH to about 5. Themixture is then concentrated in vacuo to a small volume, diluted withwater and extracted with chloroform. The extract is washed with water,dried and concentrated. The concentrate is chromatographed overacid-washed alumina and eluted with ether-petroleum ether mixtures togive 17a chloroethynyl-19-nor-5-androstene-3B,17,3-diol 17-acetate.

Ten grams of 17a chloroethynyl-l9-nor-5-androstene-35,17B-diol-l7-acetate in 150 ml. of tetrahydrofuran is treated with ml.of 0.9 m. monoperphthalic acid in ethyl acetate and allowed to standovernight at room temperature. The reaction mixture is diluted withethyl acetate and washed sequentially with aqueous sodium bicar bonate,sodium bisulfite, and sodium bicarbonate solution. The organic layer isdried and concentrated in vacuo. The product is fractionallycrystallized from aqueous ethanol to yield the17a-ch1oroethynyl-19-nor-androstane-3 3-,17B-diol-5,6ot-oxide-17-acetate.

Five grams of 17 chloroethynyL19-nor-androstane-3B,l7fi-diOl-5,6oc-0Xide 17 acetate in benzene is added to a reagentprepared from 3.4 g. of magnesium, 10 ml. of methyl iodide and 45 ml. ofether. The mixture is stirred and 70 ml. of solvent is removed bydistillation. After refluxing for 5 hours, the mixture is cooled,acidified with dilute hydrochloric acid and the organic layer washed toneutrality. The organic phase is dried and concentrated in vacuo.Crystallization from aqueous ethanol gives the 6 8 methyl 17ozchloroethynyl-19-nor-androstane-3B, 5a,l7,8-triol.

The oxdizing reagent is prepared by diluting 2.7 g. of C 0 and 2.3 ml.of concentrated sulfuric acid to 10 ml. with water. A soution of 1.90 g.of 6fl-methyl-17a-chloroethynyl 19 nor-androstane-6/3,5a,17,8-triol in300 ml. of acetone is cooled to 0 and treated with 1.85 ml. of theoxidizing reagent. The solution is diluted with ice water and extractedwith ether. The ether extract is washed sequentially with Water, andaqueous sodium bicarbonate and then dried and concentrated in vacuo, togive 613- methyl 17cc chloroethynyl-19-nor-androstane-5a,17pdiol-3-one.The crude product, dissolved in ml. of methanol, 75 ml. of 1 N sodiumhydroxide is added and the mixture is stirred at room temperature undernitrogen for 24 hours. The mixture is concentrated to half volume runder vacuo at 25 C. and then poured into ice water.

The product is extracted with ether. The ether layer is washed withwater, dried and concentrated. The concentrate is chromatographed onacid-washed alumina and the product eluted with ether-petroleum ethermixture to yield 60: methyl 17oz chloroethynyl-19-nor-androstene-17B-ol-3-one.

In accordance with the above procedures, but starting with thel7a-bromoethynyl-19-nor-4-androstene-1718-01-3- one in place of the1fiat-chloroethynyl-19-nor-4-androstene-l7fi-ol-3-one there is obtainedas intermediates, the 17a-bromoethyny1-l9-nor 3,5 androstadiene-3,l7/3-diol diacetate, 17a-bromoethynyl-19-nor-5-androstene-3B, 17,8-diol,17u-bromoethynyl 19 nor-androstane-3j3,17fidiol-5,6a-oxide-17-acetate,and the 6,8-methyl-17a-bromoethynyl-l9-nor-androstane-3 8,5a,l7fl-triol,and as product, theoat-methyl-17a-br0m0ethynyl-19-nor-4-androstene-l7fi-01-3-one.

EXAMPLE 32 100 mg. of 6a-methyl-17a-chloroethynyl-19-nor-4-androstene-17B-ol-3-one is heated with 1 cc. of acetic anhydride and 1.2cc. of pyridine on the steam bath overnight. The reaction mixture isthen poured onto ice and extracted with chloroform. The extract iswashed with water and concentrated. The concentrate is chromatographedover acid-washed alumina and eluted with mixtures of ether-petroleum andether to give 6a-methyl-l7u-chloroethynyl-19 nor-4-androstene-17B-ol-3-one acetate.

In accordance with the above procedures, but starting with the6tz-methyl-17a-bromoethynyl-l9-nor-4- androstene-l7fl-ol-3-one in placeof the 6a-methyl-17otchloroethynyl-19-nor-4-androstene-17,3-01-3-one,there is obtained theGot-methyl-17a-bromoethynyl-19-nor-4-androstene-17fi-o1-3-one acetate.

A mixture of 500 mg. of theoa-methyl-lh-chloroethynyl-19-nor-4-androstene-17,9-ol-3-one, 10 ml. ofdimethylformamide, 20 ml. of methyl iodide, and 1.5 gms. of silver oxideis stirred at room temperature for 4 days. An additional /2 gm. ofsilver oxide is added at the end of each day. One hundred m1. ofchloroform is then added to the reaction mixture which is then stirredfor one hour, filtered, and the filtrate is evaporated to dryness. Theresidual oil is chromatographed with acidwashed alumina and eluted withmixtures of ether and petroleum ether to give6ot-l'l'lethyl-170L-ChlOIO6thyIlyl-19-nor-4-androstene-17B-methoxy-3-one.

In accordance with the above procedures, but starting with the6a-methyl-17a-bromoethynyl-l9-nor-4-androstene-l7 3-ol-3-one in place ofthe 6a-methyl-l7m-chloroethynyl-l9-nor-4-androstene-17,3-ol3one, thereis obtained the6a-methyl-17a-bromoethynyl-19-nor-4-androstene-17/8-methoxy-3-one.

EXAMPLE 33 To a solution of 200 mg. of the 17a-chloroethynyl-19-nor-3,5-androstadiene-3,17B-diol diacetate (Example 17) dissolved in 2ml. of acetic acid is added, with stirring, 56 ml. ofN-chlorosuccinimide and 2 ml. of tetrahydrofuran containing 5% of dryHCl. The reaction is stirred at room temperature for 2 /2 hours. Sodiumbicarbonate is added. The reaction mixture is then extracted with ether.The ether extracts are dried and concentrated. The material ischromatographed over acid-washed alumina, and eluted withether-petroleum ether mixtures to give6a-chloro-17a-chloroethynyll9-nor-androstenel7 8-ol-3-one acetate.

In accordance with the above procedures, but startling with thel7u-bromoethynyl-l9-nor-3,5-androstadiene-3, 17fi-diol diacetate inplace of the 17a-chloroethynyl-19- nor-3,5-androstadiene-3,17B-dioldiacetate there is obtained theout-chloro-17a-bromoethyny1-19-nor-androstene-l7B-ol-3-one acetate.

30 EXAMPLE 34 A solution of the1fiat-chloroethynyl-l9-nor-3,S-androstadiene-3,l7fl-diol diacetate inaqueous tetrahydrofuran is treated with a slow stream of perchlorylfluoride for one hour and then allowed to stand at room temperature foran additional 2 hours. The reaction mixture is poured on ice andextracted with chloroform. The chloroform layer is washed with aqueousbicarbonate solution, dried and concentrated in vacuo. The crudematerial is dissolved in acetic acid saturated with gaseoushydrogenchloride and allowed to stand for one hour at room temperature. Thereaction mixture is poured on ice and extracted with chloroform. Thechloroform layer is washed with aqueous bicarbonate solution, dried andconcentrated in vacuo. The concentrate is chromatographed on acid-washedalumina to yield the 6a-fiuoro-l7a-chloroethynyl-19-nor-androstene-l7B-ol-3-one acetate.

In accordance with the above procedures, but starting with thel7a-bromoethynyl-19-nor-3,5-androstadiene-3, 17fi-diol diacetate inplace of the 17a-chloroethynyl-19- nor-3,5-androstadiene-3,17,8-dioldiacetate there is obtained the 6OL-fluOl'O 17ccbromoethynyl-19-nor-androstene-17B-o1-3-one acetate.

A mixture of 500 mg. of the 17a-chloroethynyl19-nor-4-androstene-17,8-ol-3-one (Example 3) 10 ml. of dimethylformamide, 20ml. of methyl iodide, and 1.5 gm. of silver oxide is stirred at roomtemperature for 4 days. An additional /2 gm. of silver oxide is added atthe end of each day. One hundred ml. of chloroform is then added to thereaction mixture which is then stirred for one hour, filtered, and thefiltrate is evaporated to dryness. The residual oil is chromatographedwith acid-washed alumina and eluted with mixtures of ether and petroleumether to give 17 a chloroethynyl 19 nor 4-androstene-17flmethoxy-3-one.

In accordance with the above procedures, but starting with the17u-bromoethynyl-l9-nor-4-androstene-17,8-01-3- one in place of the17a-chloroethynyl-19-nor-4-androstene-17 3-0l-3-One there is obtainedthe 17u-bromoethynyl-19-n0r-4-androstene-17fl-methoxy-3-one.

A mixture of 1 g. ofHot-chloroethynyl-19-nor-4-androstene-17fi-methoxy-3-one, 10 ml. ofacetic anhydride and mg. of p-toluenesulfonic acid is heated on thesteam bath for one hour and allowed to stand overnight at roomtemperature. It is then poured into ice water and left to stand at roomtemperature for one half hour. The reaction mixture is then extractedwith ether. The ether extract is washed with water and aqueous sodiumbicarbonate, dried, and concentrated to yield the crude 17ccchloroethynyl 19 nor 3,5 androstadiene 17B- methoxy-3-ol acetate.

In accordance with the above procedures, but starting with the 17ozbromoethynyl 19 nor-4-androstene-17B- methoxy-3-one in place of the17a-ch1oroethyny1-19-nor- 4-androstene-17,8-methoxy-3-one, there isobtained the 17m bromoethynyl 19 nor 3,5 androstadiene 1715-methoxy-3-ol acetate.

To a solution of 200 mg. of the 17a-chloroethynyl-19-nor-3,S-androstadiene-l7/3-methoxy3-ol acetate dissolved in 2 ml. ofacetic acid is added, with stirring, 56 ml. of N-chlorosuccinimide and 2m1. of tetrahydrofuran containing 5% of dry HCl. The reaction is stirredat room temperature for 2 /2 hours. Sodium bicarbonate is added. Thereaction mixture is then extracted with ether. The ether extracts aredried and concentrated. The material is chromatographed over acid-washedalumina, and eluted with ether-petroleum ether mixtures to giveGot-chloro- 17a chloroethynyl 19 nor 4 androstene 1713- methoxy-3-one.

In accordance with the above procedures, but starting with the17u-bromoethynyl-19-nor-3,S-androstadiene-17 3- met-hoxy-3-ol acetate inplace of the Hot-chloroethynyl-19-nor-3,S-androstadiene-17fi-methoxy-3-ol acetate there is obtained the6a-chloro-17wbromoethynyl-19-nor-4-androstene-17fl-methoxy-3-one.

31 EXAMPLE 3s A solution of the17a-chloroethynyl-l9-nor-3,S-androstadiene-17pt-methoxy-3-ol acetate inaqueous tetrahydrofuran is treated with a slow stream of perchlorylfluoride for one hour and then allowed to stand at room temperature foran additional 2 hours. The reaction mixture is poured on ice andextracted with chloroform. The chloroform layer is washed with aqueousbicarbonate solution, dried and concentrated in vacuo. The crudematerial is dissolved in acetic acid saturated with gaseous hydrogenchloride and allowed to stand for one hour at room temperature. Thereaction mixture is poured on ice and extracted with chloroform. Thechloroform layer is washed with aqueous bicarbonate solution, dried andconcentrated in vacuo. The concentrate is chromatographed on acidwashedalumina to yield the 60t-fll1010-170t-Chl0106l'hYHyl-19-nor-4-androstene-17p-methoXy-3-one.

In accordance with the above procedures, but starting with the17a-bromoethynyl-19-nor-3,S-androstadiene-li'fimethoxy-3-ol acetate inplace of the 17a-chloroethynyl- 19 nor 3,5 androstadiene 17,8 methoxy 3ol actate, there is obtained the6a-fluoro-17ot-bromoethynyll9-nor-4-androstene-17 3-methoxy-3-one.

EXAMPLE 36 A solution of 100 mg. of the 17a-chloroethynyl-l9-nor-4-androstene-17,8-ol-3-one and 50 mg. of Lindlar catalyst in 10 cc. ofethyl acetate is treated with hydrogen until one mole of hydrogen hasbeen absorbed. The mixture is filtered and concentrated to yield thecrude chlorovinyl compound. Chromatography yields the pure product 21-chloro-19-nor-4,20-pregnadiene-17B-ol-3-one.

In accordance with the above procedure, but starting with thel7a-bromoethynyl-l9-nor-4-androstene-175-01-3- one, the17a-chloroethynyl-l9-nor-5 (10) -ar1drostene-17fiol3-one, or the17a-bromoethynyl-19-nor-5 (10)-androstene-17B-ol-3-one, in place of the17u-chloroethynyl-19- nor-4-androstene-17fl-ol-3-one, there is obtainedthe 21- bromo-l9-nor-4,20-pregnadiene-17fi-ol-3-one, 2l-chloro-19-nor-5(10),ZO-pregnadiene-l7B-0l-3-one, or 21-bromo- 19-nor-510),ZO-pregnadiene-l7/3-ol-3-one respectively.

EXAMPLE 37 A suspension of platinum oxide in 10 cc. of ethanol isreduced and 100 mg. ofl7a-chloroethynyl-19-nor-4-androstene-l7fi3-ol-3-one is added. Reductionproceeds until two moles of hydrogen have been absorbed. The solution isfiltered, concentrated and chromatographed on alumina to yield thecorresponding 17a-chloroethyl derivative (21-chloro-19-nor-4-pregnene-17fi-ol-3-one) In accordance with the aboveprocedure, but starting with the17a-bromoethynyl-19-nor-4-androstene-17fl-ol-3- one, the17a-chloroethynyl-5(IO-androstene-l7fi-ol-3-one, or thel7u-bromoethynyl-5(10)-androstene-17fl-ol-3-one in place of the 17a-chloro-ethynyl-19-nor-4-androstene- 17 3-0l-3-one, there is obtainedthe 2l-bromo 19-nor-4- pregnene-17fi-ol-3-one, the 21-chloro-5( 10)-pregnene-17[3- ol-3-one, or the 21-brorno-19-nor-pregnene-17B-ol-3-onerespectively.

Various changes and modifications may be made in carrying out thepresent invention without departing from the spirit and scope thereof.

We claim:

1. Compounds selected from the group consisting of 17a haloethynyl 4androstene 17B ol 3 one and the 17,8-lower alkyl ethers and 17/3-loweralkanoyl esters thereof.

2. Compounds selected from the group consisting of 1701. haloethynyl-1,4 androstadiene 17B ol 3 one and the 17fi-lower alkyl ethers and17fi-lower alkanoyl esters thereof.

3. Process for the preparation of 17a-haloethynyl-4-androstene-17fi-ol-3-one which comprises reacting 17ahaloethynylandrostene 313,175 diol with aluminum isopropoxide and acetone.

wherein the broken lines between carbon atoms 4-5, 56 and 5-10 indicatethat a double bond may be present in these positions and wherein R isselected from the group consisting of hydrogen, lower alkyl and loweralkanoyl, X is selected from the group consisting of --CECC1, -CH=CHC1,-C=. =C'BI, CH=OHBr, -CH CH Cl and -CH CH Br, and Y is selected fromthegroup consisting of hydrogen, methyl, chloro and fluoro.

5. Compounds selected from the group consisting of haloethynyl 19 nor 4androstene 17B ol 3- one and the 17,8-lower alkyl ethers and 17,8-loweralkanoyl esters thereof.

6. Compounds selected from the group consisting of 17a haloethynyl 19nor S androstene ol- 3-one and the 17B-lower alkyl ethers and 17,8-loweralkanoyl esters thereof.

7. Compounds selected from the group consisting of Her-haloethynyl 19-nor 5(10) androstene 17B- ol-3-one and the 17fi-lower alkyl ethers andUri-lower alkanoyl esters thereof.

8. Compounds selected from the group consisting of 6a methyl 17ahaloethynyl 19 nor 4 androstene- 17/8-ol-3-one and 17,8-lower alkylethers and 17fi-lower alkanoyl esters thereof.

9. Compounds selected from the group consisting of 6a halo 17ozhaloethynyl 19 nor 4 androstene- 17,8-ol-3-one and l7i3-lower alkylethers and 17fi-lower al-kanoyl esters thereof.

10. A compound of the formula:

wherein R is selected from the group consisting of hydrogen and methyl.

11. A compound of the formula:

OH CH:

12. A compound of the formula:

: --GECCl 13. A compound of the formula:

34 14. 17a chloroethynyl 19 nor 4 -androstene- 17,8-methoxy-3-one.

15. 170: chloroethynyl 19 nor 4 androstene- 1718-n-propoxy-3-one.

16. 170a chloroethynyl 19 nor 5(10) androstene- 17/3-methoxy-3-one.

No references cited.

ELBERT L. ROBERTS, Primary Examlner 1O U.S. C1. X.R.

